VALIDATION OF ELECTROSTATIC SPRAY AS A LOW-VOLUME ... · Piansay, Sherre Chambliss, and Yanjie...

VALIDATION OF ELECTROSTATIC SPRAY AS A LOW-VOLUME SANITIZATION

METHOD FOR FOOD PROCESSING SURFACES

By

SHAWN MATTHEW LYONS

(Under the direction of Mark A. Harrison and S. Edward Law)

ABSTRACT

In this study, air-assisted, induction-charged sprays of sanitizers were applied to

inoculated food contact surfaces to evaluate their ability to reduce populations of Salmonella.

Electrostatically charged sprays (-7.2 mC/kg charge-to-mass ratio) deposited more active

ingredient and carrier liquid onto target surfaces than uncharged sprays from the same nozzle and

a conventional hydraulic nozzle (p<0.05). Charged sprays at lowered biocide rates reduced

Salmonella population on target surfaces greater than or equal to hydraulic sprays with full-rate

biocide for 8 of 9 surface and orientation combinations evaluated (p<0.05). Peracetic acid sprays

were more effective than quaternary ammonium compound sprays from all nozzles in 9 of 9

surface and orientation combinations evaluated (p<0.05).

Index words: electrostatic spray, quaternary ammonium compounds, peracetic acid, food contact

surfaces. sanitizers, Salmonella



By

SHAWN MATTHEW LYONS

B.S., Clemson University, 2008

A Thesis Submitted to the Graduate Faculty of the University of Georgia in Partial Fulfillment

of the Requirements for the Degree

MASTER OF SCIENCE

ATHENS, GEORGIA

2011

© 2011

Shawn Matthew Lyons

All Rights Reserved



by

SHAWN MATTHEW LYONS

Major Professors: Mark A. Harrison

S. Edward Law

Committee: Faith J. Critzer

William C. Hurst

Electronic Version Approved:

Maureen Grasso

Dean of the Graduate School

The University of Georgia

May 2010

iv

ACKNOWLEDGEMENTS

I would like to thank my major professors Dr. Mark Harrison and Dr. Edward Law for

their guidance, technical expertise, and support. I also thank Dr. William Hurst and Dr. Faith

Critzer for serving on my committee.

I thank Ruth Ann Morrow for laboratory training and support. I would like to thank my

colleagues who assisted the project: Chi-ching Lee, Amudhan Ponrajan, Winnie Lim, Belle

Piansay, Sherre Chambliss, and Yanjie Tang.

I would also like to thank Mr. Pat Harrell and Mr. Danny Morris for their assistance in

the fabrication of the spray setup used for the project.

Lastly, I would like to thank National Institute of Food and Agriculture, USDA, under

special project #2009-51110-20161 and the Georgia Agricultural Experiment Stations for

funding this project.

v

TABLE OF CONTENTS

Page

ACKNOWLEDGEMENTS .......................................................................................................iv

LIST OF TABLES .....................................................................................................................vi

LIST OF FIGURES .................................................................................................................... x

CHAPTER

1. INTRODUCTION ..........................................................................................................1

2. LITERATURE REVIEW ...............................................................................................3

References ........................................................................................................ 17

3. VALIDATION OF ELECTROSTATIC SPRAY AS A LOW-VOLUME

SANITIZATION METHOD FOR FOOD PROCESSING SURFACES1 ..................... 23

Abstract ............................................................................................................ 24

Introduction and Literature Review ................................................................ 25

Materials and Methods .................................................................................... 28

Results and Discussion ..................................................................................... 38

Acknowledgements........................................................................................... 45

References ........................................................................................................ 46

4. CONCLUSION ............................................................................................................ 50

5. APPENDIX A .............................................................................................................. 73

vi

LIST OF TABLES

Page

Table 3.1. Tracer deposition on stainless steel coupons normalized to equal tracer mass

dispensed toward target coupons using three spray treatments (Hydraulic:

hydraulic nozzle dispensing 600 ml of sanitizer spray/min, Uncharged: uncharged

electrostatic nozzle dispensing 100 ml of sanitizer spray/min, Charged: charged

electrostatic nozzle dispensing 100 ml of sanitizer spray/min). Front side, back

side: n = 12. Side: n = 24.

……………………………………………………………………………………51

Table 3.2. Carrier liquid deposition on stainless steel coupons using three spray treatments

(Hydraulic: hydraulic nozzle dispensing 600 ml of sanitizer/min, Uncharged:

uncharged electrostatic nozzle dispensing 100 ml of sanitizer/min, Charged:

charged electrostatic nozzle dispensing 100 ml of sanitizer/min). Front side, back

side: n = 12. Side: n = 24.

……………………………………………………………………………………52

Table 3.3. Reduction of Salmonella populations on the front surfaces of stainless steel

coupons after treatment with peracetic acid, quaternary ammonium compounds,

and tap water using three spray treatments. Biocide dispensed full-rate hydraulic

vs. equivalence-rate charge and uncharged. n = 12. ............................................ 55

Table 3.4. Reduction of Salmonella populations on the back surfaces of stainless steel


and tap water using three spray treatments (Hydraulic: hydraulic nozzle

vii

dispensing 600 ml of sanitizer spray/min, Uncharged: uncharged electrostatic

nozzle dispensing 100 ml of sanitizer spray/min, Charged: charged electrostatic

nozzle dispensing 100 ml of sanitizer spray/min). Biocide dispensed full-rate

hydraulic vs. equivalence-rate charge and uncharged. n = 12. ............................. 57

Table 3.5. Reduction of Salmonella populations on the side surfaces of stainless steel







Table 3.6. Reduction of Salmonella populations on the front surfaces of polyvinyl chloride






hydraulic vs. equivalence-rate charge and uncharged. n =12. .............................. 61

Table 3.7. Reduction of Salmonella populations on the back surfaces of polyvinyl chloride


and tap water using three spray treatments1 (Hydraulic: hydraulic nozzle



viii



Table 3.8. Reduction of Salmonella populations on the side surfaces of polyvinyl chloride







Table 3.9. Reduction of Salmonella populations on the front surfaces of waxed cardboard







Table 3.10. Reduction of Salmonella populations on the back surfaces of waxed cardboard







ix

Table 3.11. Reduction of Salmonella populations on the side surfaces of waxed cardboard







Table A.1. Calculated spray interception by target coupon………………………………….79

Table A.2. Sanitizer mixing volumes for equivalence point test concentrations…………….87

Table A.3. Sanitizer mixing volumes for microbiological assessment of varying surfaces and

target orientations………………………………………………………………...90

x

LIST OF FIGURES

Page

Figure 3.1. Effect of relative rate of peracetic acid sanitizer on population density of

Salmonella on stainless steel coupons (20.7 cm2) applied as a low volume

electrostatically charged spray. Values are means and standard errors of three

replicate experiments each with four sample coupons. The regression equation is

y = 1.347ln(x) + 4.458, (R2 = 0.77). The dashed line and open symbol with error

bars represent the mean log CFU reduction per coupon face when peracetic acid is

applied as a hydraulic spray at full rate. n = 12. .................................................. 53

Figure 3.2. Effect of relative rate of quaternary ammonium sanitizer on population density of

Salmonella on stainless steel coupons (20.7 cm2) applied as a low-volume



y = 1.934x + 0.901, (R2 = 0.99). The dashed line and open symbol with error bars

represent the mean log CFU reduction per coupon face when peracetic acid is

applied as a hydraulic spray at full rate. n = 12. .................................................. 54

Figure 3.3. Population reduction of Salmonella on the front surface of stainless steel coupons

after treatment with peracetic acid, quaternary ammonium compounds, and tap

water using three spray treatments (Hydraulic: hydraulic nozzle dispensing 600

ml of sanitizer spray/min, Uncharged: uncharged electrostatic nozzle dispensing

100 ml of sanitizer spray/min, Charged: charged electrostatic nozzle dispensing

100 ml of sanitizer spray/min). Biocide dispensed full-rate hydraulic vs.

xi

equivalence-rate charge and uncharged. Initial population: 106 CFU/coupon face.

Detection limit: 1.9 log CFU/coupon face. n = 12............................................... 56

Figure 3.4. Population reduction of Salmonella on the back surface of stainless steel coupons








Figure 3.5. Population reduction of Salmonella on the side surfaces of stainless steel coupons








Figure 3.6. Population reduction of Salmonella on the front surfaces of polyvinyl chloride






xii

hydraulic vs. equivalence-rate charge and uncharged. Initial population: 106

CFU/coupon face. Detection limit: 1.9 log CFU/coupon face. n = 12. ................ 62

Figure 3.7. Population reduction of Salmonella on the back surfaces of polyvinyl chloride








Figure 3.8. Population reduction of Salmonella on the side surfaces of polyvinyl chloride








Figure 3.9. Population reduction of Salmonella on the front surfaces of waxed cardboard






xiii



Figure 3.10. Population reduction of Salmonella on the back surfaces of waxed cardboard








Figure 3.11. Population reduction of Salmonella on the side surfaces of waxed cardboard








Figure A.1. Food Science and Technology spray apparatus and target chamber – side

view……………………………………………………………………...……….74

Figure A.2. Food Science and Technology spray target chamber – frontal

view………………………………………………………………………………75

Figure A.3. Spray arm pathway dimensions…………………………………………….……76

xiv

Figure A.4. Input voltage source: Lambda Model LLS6108 vs. output volts measured by:

Extech Digital Multimeter Model EX410………………………………………..83

Figure A.5. Fluorescent tracer deposition on stainless steel coupons………………………...85

Figure A.6. Spot inoculation of target coupons in BSL2 biosafety cabinet…………………..91

Figure A.7. Spot inoculated waxed cardboard target coupons………………………………..92

Figure A.8. Spot inoculated stainless steel target coupons…………………………………...93

Figure A.9. Spot inoculated PVC target coupons…………………………………………….94

Figure A.10. Target coupon orientation with respect to incoming spray vector……………....95

1

Chapter 1

INTRODUCTION

This study evaluated decontamination efficacy of three types of spraying systems by

applying sanitizers (biocide + carrier liquid) to food contact surfaces inoculated with Salmonella

enterica. Decontamination of food contact surfaces is a key hurdle in the prevention of buildup

and transfer of pathogenic organisms to foods and in turn to consumers. Efficient application

methods deposit sanitizers onto intended target surfaces and reduce the amount of off-target

losses.

Four specific objectives were met:

1. Design and construct a system in which spray targets could be consistently sprayed using a

repeatable robotic arm.

2. Quantify the spray deposition density of active ingredient onto target coupons using an

idealized stainless steel surface in three orientations, front side, back side, and side, with

three spray nozzle treatments:

a. Charged droplets produced by pneumatic atomizing spray nozzle

b. Uncharged droplets produced by pneumatic atomizing spray nozzle

c. Spray droplets produced by a conventional hydraulic-atomizing spray nozzle.

3. Identify an equivalence point at which a reduced mass of active ingredient dispensed from

an air-assisted electrostatic nozzle with charging on has the same effect in terms of

population reduction of Salmonella as a hydraulic-atomizing spray nozzle dispensing full

rate mass of active ingredient for stainless steel coupons.

2

4. Using equivalence points from objective 3, evaluate each nozzle treatment for efficacy in

terms of population reduction of Salmonella using two liquid sanitizers, applied to three

food contact surfaces, in three target orientations.

3

Chapter 2

LITERATURE REVIEW

Electrostatic Principles and Nozzle Design

Electric force fields imparted on small particles allow them to be managed into many

applications such as xerography, sandpaper manufacture, textile flocking, discrete-droplet

electrostatic printing, and agricultural pesticide application (41). Law developed an embedded-

electrode electrostatic-induction spray-charging nozzle for use in deposition of liquid pesticide

droplets having a low to middle range resistivity of 10-1

to 104 ohm m onto agricultural plants

(41). A positive potential is applied to a cylindrical electrode which has a grounded liquid

passing through it which accumulates an induced negative charge (41). The addition of a

pneumatic force of as low as 30 psi shears this liquid into a film which ruptures into negatively

charged droplets while pushing the spray liquid toward the desired target (41). Volume median

diameter produced by the developed nozzle was approximately 30 μm (41). Evaporation of

small diameter aqueous sprays in warm ambient temperatures was initially thought to be a

challenge to electrostatic spray applications, but rapid trajectory of charged droplets pushed forth

by pneumatic energy effectively reach targets before charge loss occurs (51). The

aforementioned nozzle settings initially yielded an average droplet charge-to-mass ratio of 4.8

mC/kg.

Factors that affect spray deposition include: charge-to-mass ratio, applied voltage,

atomizing air pressure, liquid flow rates, and liquid resistivity (26, 41). Charge-to-mass ratio is

the most significant operating characteristic for deposition of charged sprays. Typical charge-to-

4

mass ratio values for charged sprays of conductive liquids producing 30-50 μm volume median

diameter droplets are typically – 10 mC/kg, and 5-15 mC/kg charge-to-mass for most other

electrostatic-based applications (42, 43). An increased charge-to-mass ratio results in increased

deposition and ―wrap-around‖ effect. Law‘s embedded-electrode electrostatic-induction spray-

charging nozzle exhibited a linear relationship of increased spray-cloud current as charging

voltage increases given a fixed liquid volumetric flow rate (41). The curve became nonlinear at

approximately 1.5 to 2 kV, showing minimal increase in spray cloud current with increased

voltage applied to the induction electrode (41). Liquid flow rate was shown to have a positive

linear relationship with spray cloud current up to 80 ml/min for the developed embedded

electrode nozzle (41). Spray liquid flow rates of 30, 45, and 60 mL/min (lower flow rates) were

found to have better chargeability than 250, 450, and 600 mL/min (higher flow rates) (50).

Liquid resistivity was seen to affect charge-transfer at levels greater 104 ohm m (41). Surfactants

were observed to increase the charge on spray droplets significantly (59). Thirty to forty psi of

compressed air was sufficient to atomize spray liquid into the small droplets required for Law‘s

embedded electrode design (41).

Electrostatic Spray Mass Transfer and Application

Fluorometric analysis of tracer spray deposits on target surfaces has been a common

method of measuring performance of sprayers on variety of surface types and morphologies (24).

Mass of tracer within a spray liquid deposited is used to calculate the advantage of electrostatic

spray deposition over uncharged and conventional sprays. In addition to fluorometric analysis to

document mass transferred from nozzle to target, the activity of the biocontrol agent applied is

evaluated for efficacy.

5

Air-assisted electrostatic spraying technology is used in agricultural applications to

penetrate the plant canopy and efficiently deliver the active ingredient applied to its target area.

In cotton plants, Law showed that air-assisted charged spray deposited tracer onto leaf

undersides 1.9-fold and 2.5-fold more than air-assisted uncharged spray and hydraulic spray

methods respectively (17). Maski and Durairaj constructed a spraying system which sprayed

tracer liquid downward into simulated cotton plants made of aluminum leaves to show the effects

of charging voltage, application speed, target height, and orientation upon charged spray

deposition on leaf abaxial and adaxial surfaces. Abaxial deposition was found to be nearly zero

for uncharged sprays, while charging increased the deposition. Additionally, deposition on

adaxial surfaces was found to be greatest at 0° from horizontal. A medium nozzle ground

application speed, 0.417 m/s was found to enhance deposition on abaxial surfaces for all target

heights (49). Charged full rate application had a larger mean deposition than both uncharged and

conventional applications of whitefly pesticide mixed with dye tracer (ng/cm2) on cotton leaves

in top- and mid-canopy locations averaged over six aerial sprays. Accordingly, electrostatic

spray charging system using a spray rate of 4.68 L/ha reduced whitefly counts on a season-long

basis to a level comparable with that of conventional spray applications of 46.8 L/ha (40). Kang

et al. found electrostatic orchard sprayers to have 4.3 times the covering area ratio than

conventional spraying systems (34). At an average charge-to-mass ratio of – 7.8 mC/kg, air-

assisted electrostatic spray showed a 4.5-fold increase in deposition of colony forming units of B.

subtilis biofungicide to control mummy berry disease on blueberry flower stigmas over that

deposited by conventional spraying (66).

Air-assisted electrostatically charged pollen sprays (charge-to-mass ratio -12 mC/kg)

deposited 5.6-fold pollen onto target germination wells on average across various target

6

orientations. Target wells parallel to the spray‘s air carrier stream showed the greatest increase

of pollen deposition for charged vs. uncharged sprays at 12.1-fold (46). Deposition of tracer

mimicking pesticide application on foliar targets of broccoli, corn, cotton, and cabbage increased

for charged sprays when compared to uncharged sprays 1.8-, 2.0-, 2.5-, and 7.0-fold respectively.

Charged spray was shown to increase deposition over conventional spraying 1.9-, 2.9-, and 4.4-

fold for broccoli, cabbage, and corn plants (45). Chaim et al. showed that pesticides applied as

charged sprays onto spherical targets had a deposition density of 62% of tracer spray liquid

dispensed at 4 mC/kg charge-to-mass ratio whereas uncharged sprays deposited only 18% (13).

Air-assisted electrostatic spray has also been evaluated for enhanced deposition of

chemical to agricultural commodities postharvest. Tracer deposition showed 1.5 to 3.4-fold

increase for charged spray vs. uncharged sprays onto metal target coupons placed in various

positions on banana bunches (43). Tracer deposition showed 1.9 to 2.5-fold increase for charged

spray (charge-to-mass 5-10 mC/kg) vs. uncharged sprays onto plastic target coupons placed in

various positions on banana bunches (43). Due to increased spray deposition of fungicide,

banana spoilage in shipment was controlled showing 86% of electrostatically treated bananas

crown-rot free versus 74% and 36% for conventionally treated and control treatments while

using half of the active ingredient chemical (43). Kim et al. showed that charged sprays had

greater mass deposition and uniformity of deposition than uncharged sprays on spherical targets

like tomatoes, and apples (36).

Air-assisted electrostatic sprays are also used for application of sanitizers in efforts to

decontaminate surfaces containing human pathogens. In an evaluation of decontamination of

humans in the event of a bio-terrorism attack, Law et al. found tracer spray with a charge-to-

mass ratio of -16 mC/kg provided 1.8-fold increased deposition over uncharged sprays as

7

averaged over 52 sites on grounded mannequin and human subjects. Microbiological assessment

in the same system showed a charged spray of quaternary ammonium onto Pseudomonas

inoculated targets placed around body sites to have an average 2.38 log CFU reduction compared

to uncharged sprays with 0.66 log CFU reduction (44). Similarly, Cooper and Law evaluated

electrostatically charged sprays for sunless tanning of the human body, finding charged spray

with a charge-to-mass ratio of -13 mC/kg had an average two-fold greater deposition than

uncharged spray on stainless steel target coupons placed around grounded mannequins and

human subjects. In those applications, the respirable mist was peaked at 40 mg/m3 for charged

sprays and 150 mg/m3 for uncharged sprays (14). Hsu et al. applied electrolyzed water via

electrostatic spray gun to produce a 3-4 log CFU reduction of Listeria monocytogenes on

surfaces (32). Russell applied electrolyzed water to unhatched eggs inoculated with Salmonella

to produce at least a 4 log CFU reduction over the control treatment (62). In recent years, hand-

held electrostatic spraying equipment has been used in efforts to disinfect schools, hospitals, and

hotels (67). Handheld sprayers were used in 2009 to apply hospital grade disinfectant on hand-

contact surfaces in the Washington D.C. Metro bus system in efforts to control H1N1 influenza

virus (70).

Sanitation

Sanitation is defined as ―the application of a science to provide wholesome food

processed, prepared, merchandised, and sold in a clean environment by healthy workers; to

prevent contamination with microorganisms that cause foodborne illness; and to minimize the

proliferation of food spoilage organisms‖ (35). Four major inputs are associated with sanitation

processes: mechanical or kinetic energy, chemical energy, thermal energy, and time (47).

Cleaning and sanitizing food processing surfaces can be divided into several stages: wetting and

8

penetration of the surfaces with water and cleaning solution, the reaction of the cleaning solution

with the soil and surface, the prevention of re-deposition back onto the cleansed surface, the

wetting by the sanitizing solution of residual microorganisms to produce a biocidal or biostatic

action, and dispersion of microorganisms if needed (47). There are three main cleaning

procedures: manual application, immersion/clean-out-of-place (COP), and clean-in-place (CIP)

(63).

Out of ten industries evaluated in the Netherlands, the food industry used the third most

water, behind chemical and refinery industries (8).Cleaning and sanitizing a food processing

plant can represent a large portion of water used in production depending upon the product. For

example, the percentages of water used in beverage, dairy, vegetable, and meat processing were

25, 49, 15, and 48%, respectively (61).

Surface and liquid characteristics play a large role in the ability for an applied liquid to

cover an intended target surface. Surface roughness and wettability were found to have a

synergistic effect on fouling cleaning and disinfection. Durr showed double linear regressions

based on both independent variables had coefficients of correlation 0.99 (21). Stainless steel is

comparable in its biological cleanability to glass, and significantly better than polymers,

aluminum or copper (5).

Surface energy is measured using contact angles which describe how a liquid wets a

solid. The shape of a drop of liquid on a solid surface allows conclusions to be drawn

concerning the forces acting at the interface and the work of adhesion (19). Large contact angles

indicate a hydrophobic surface (wetting) while small contact angles indicate a hydrophilic

surface (wetting). Through dynamic contact angle analysis, Davies et al. showed that clean

stainless steel exhibited hydrophilic characteristics while polymers polyphenylene (PPE),

9

polytetrafluoroethylene (PTFE), ethylene propylene diene monomer (EPDM), and silicone

tubing (ST) were all very hydrophobic (18). Midelet and Carpentier reported water contact

angles on clean stainless steel to be significantly lower than those on polyvinyl chloride (PVC)

conveyor belting (52).

Sanitizers

To sanitize means to adequately treat food-contact surfaces by a process that is effective

in destroying vegetative cells of microorganisms of public health significance, and in

substantially reducing numbers of other undesirable microorganisms, but without adversely

affecting the product or its safety for the consumer (4, 25). Sanitizers are categorized as no-rinse

food contact surface sanitizers and non-food-contact surface sanitizers which are tested against

recommended standard strains (48). EPA sanitizer tests for inanimate, non-food contact surfaces

require at least 99.9% bacterial population reduction over the parallel control count within 5

minutes (22). Sanitizers applied to food contact surfaces are defined as incidental food additives

under the Federal Food, Drug, and Cosmetic Act, as amended (21 U.S.C. 201 et seq.).

Acceptable results must demonstrate a 99.999% reduction in the number of microorganisms

within 30 seconds (23). Five log reductions are satisfactory results in standardized suspension

tests such as the AOAC Germicidal and Detergent Sanitizing Action of Disinfectants 960.09,

which is used for efficacy testing of non-halogen sanitizers.

Though suspension tests provide a simple repeatable way to measure the biocidal action

of sanitizers, the test has several limitations. Van Klingeren et al. observed that covering a dried

inoculum with disinfectant without any further mechanical action to improve contact between

organisms and disinfectant, will usually result in lower reduction factors than those obtained with

suspension tests (69). Kim et al. reported that biocides were most effective in the order of

10

planktonic cells > cells inoculated and dried onto stainless steel > cells in biofilms on stainless

steel (37). Moretro et al. observed that recommended standard test strains for sanitizer efficacy

testing were more sensitive to sanitizers than were persistent strains isolated from fish feed

factories (55). Due to the limitations of suspension testing, Gibson et al. suggested that a three

phase approach be done: 1) suspension testing to establish minimum standards for bactericidal

effect; 2) suspension tests and surface tests which simulate practical conditions; and 3) a ‗field

trial‘ type of assessment (27).

A biocide is a substance that provides microbial control of a process (48). Commonly

used biocides in the food industry are chlorine compounds, quaternary ammonium compounds

(quats), amphoterics, iodine compounds, peracetic acid, and acid anionic compounds (47).

Biocide choice for the purpose of sanitizing a surface depends on the intended application. Some

physical and chemical factors that determine the efficacy of biocides are: exposure time,

temperature, concentration, equipment cleanliness, pH, water hardness, and microbial population

(48). Longer exposure of a bacterial population to biocide produces a greater population

reduction. The ideal temperature for biocidal sanitizer application is 21 to 38°C as some

biocides vaporize at high temperatures (48). Water hardness negatively affects some sanitizers

such as quats when calcium and magnesium salts are over 200 ppm. Ideally, water used for

cleaning and sanitizing is soft, or 0-60 ppm (33). A high number of bacteria initially present on a

surface results in a higher number of possible survivors to a biocidal treatment.

Desired biocide properties include: exhibit broad spectrum inactivation of bacteria,

yeasts, and molds; ability to work in presence of organic matter and in varying water hardness,

pH and temperature conditions; be non-toxic, non-irritating, odorless, highly water soluble,

11

inexpensive, and easy to measure; and be chemically stable when concentrated and in solution

(47, 48).

There are many methods in which chemical biocides suspended in water can be applied

to a surface. These methods include spray sanitizing, fogging, and flood sanitizing (48). Spray

or mist sanitizing and flood sanitizing are applied via spray nozzles, but differ in the fact that

flood sanitizing methods apply large quantities of sanitizer to ensure extensive exposure which

increases costs of sanitizer and water while simultaneously creating a wet condition (48). Spray

or mist sanitizing is the most commonly used method for applying sanitizers. The efficiency of

conventional spraying depends on (i) the water pressure employed, (ii) the volume of water used,

(iii) the water temperature, (iv) the distance of the target from the spray origin (contact force), (v)

the time of exposure of the target to the spray, (vi) the type and number of spray nozzles used,

(vii) the nozzle orientation, and (viii) the spray pattern (60). Spray or mist is usually delivered

using small hand-pumped containers, ‗knapsack‘ sprayers or pressure washing systems at low

pressure. One disadvantage of misting is that it will only ―wet‖ vertical smooth surfaces, giving

a contact time of 5 minutes or less (47). Among 18 food factories surveyed in the U.K.,

application rates of sanitizer sprays were 0.4–572 ml/m2, with a mean and median application

rate of 102 and 18.2 ml/m2, respectively (31). In the same survey, out of 117 sites 52% of sites

used a conventional hand sprayer or other type of sprayer and 28% used high pressure sprayers

including foamers out of 117 sites (31).

Quaternary ammonium compounds are generally structured as a nitrogen atom with four

alkyl groups attached making up the cation portion of the molecule which is active biocidally,

and an anion, usually chloride or bromine to form the salt (4). Quats are absorbed on the cell

surface, diffuse through the cell wall, bind and disrupt the cytoplasmic membrane. The resulting

12

release of K+ ions and cellular contents can result in cell death (4). There are many different

types of quaternary ammonium salts, many of which are combined in sanitizer formulations in

order to provide broad spectrum biocidal activity as well as to ensure biocidal activity under

various environmental conditions. Quats have the following advantages: they are colorless and

odorless, stable in presence of organic matter, non-corrosive to metals, fairly stable in hard

water, non-irritating, effective at high pH, and they have good surfactant properties and residual

activity due to stability (48, 4). Disadvantages of quat applications include their limited

effectiveness against gram negative organisms except for Salmonella and Escherichia coli,

ineffectiveness in the presence of detergents and soaps, and foaming in CIP applications (48).

Quaternary ammonium compounds can be used at concentrations up to 200 ppm on food

contact surfaces without rinsing. Environmental surfaces such as walls, floors, drains, and

overhead structures can be sanitized at a rate of 800-1000 ppm (15). Quats have been used in a

variety of food processing settings such as egg, fishing, brewing, and sugar refining industries

(4).

Peracetic acid (PAA) is the peroxide of acetic acid also known as peroxyacetic acid.

PAA causes bacterial damage through oxidation and radical formation that results in cell

membrane damage and bacterial death (4). Knight and Craven found that 3% (v/v) PAA was

most effective over hypochlorite, acid-anionic, and quat sanitizers at reducing mixed biofilm

cultures from dairy factories on floor materials (38). Advantages of PAA are: effectiveness at

low concentrations, effectiveness in presence of organic matter, no residue, stability at a wide

temperature range, non-corrosive to stainless steel, low foaming, and effectiveness over a wide

pH range. Disadvantages of PAA use are: high cost, strong odor, irritancy, and corrosiveness to

iron and some other metals (48, 4).

13

Peracetic acid is EPA registered as a non-rinse food-contact surface sanitizer when used

at the dilution specified on the label (48). PAA breaks down into acetic acid, hydrogen peroxide,

oxygen, and water, and one preparation was given clearance as an indirect food additive by FDA

in 1986 (4). Many food processing industries, such as meat and poultry processing plants,

canneries, dairies, breweries, wineries, and soft drink plants, use PAA (4). In one case, plastic

food containers were sprayed on a conveyor belt with 0.1% PAA and 20% hydrogen peroxide in

order to sanitize for reuse.

Salmonella

Salmonella spp. are facultatively anaerobic gram-negative rod-shaped bacteria belonging

to the family Enterobacteriaceae (20). Salmonella grow optimally at 37°C and are oxidase

negative and catalase positive. Salmonella enterica subspecies enterica contains 1,504 of a total

2,541 serovars (20). Salmonella infection symptoms include diarrhea, fever, and abdominal

cramps 12 to 72 hours after infection with the illness usually lasting 4 to 7 days. Most infected

persons recover without treatment, but for some the diarrhea may be so severe that

hospitalization is needed. CDC estimates that 400 people die per year from Salmonella infection

in the U.S. Those most susceptible include young children, the elderly, and the

immunocompromised (10). In 2009, USDA‘s Economic Research Service (ERS) estimated the

number of Salmonella cases due to any source as 1.4 million with an associated cost of 2.6

billion USD per year (68).

Scallan et al. estimated 38.4 million episodes of domestically acquired foodborne illness

were caused by unspecified agents, resulting in 71,878 hospitalizations and 1,686 deaths (64).

Additionally, Scallan et al. identified 31 major pathogens which were responsible for and

estimated 9.4 million episodes of foodborne illness 55,961 hospitalizations, and 1,351 deaths. Of

14

these incidences caused by specific agents, nontyphoidal Salmonella spp. were the leading cause

of bacterial foodborne illness, 11% of total. Among all pathogens, nontyphoidal Salmonella spp.

were the leading cause of hospitalizations and deaths, 35 and 28% respectively (65). According

to the Centers for Disease Control and Prevention (CDC), in 2009 a total of 17,468 laboratory-

confirmed cases of foodborne pathogen infection were identified. Salmonella species accounted

for 7,039 reported infections, having an incidence of 15.19 per 100,000 population. That year

the top three isolated Salmonella serovars were: Enteritidis, Typhimurium, and Newport (9).

Salmonella contamination in the food factory environment is due to their ubiquitous

presence in the natural environment and likely presence in some raw ingredients (3). In the past

three years multi-state outbreaks of foodborne Salmonella infection have been associated with

food items such as: alfalfa sprouts, cheesy chicken rice frozen entrée, red and black pepper,

Italian style meats, pistachios, cantaloupes, rice and wheat cereals, peanut butter and raw

produce (11).

Surface-to-food by Contact Transfer of Bacteria

Transfer of microbiological contaminants via surface-to-food by contact plays a large

role in foodborne illness (57). Once present in a food plant, pathogenic bacteria can become

established and serve as a source of cross contamination for products that enter the process line

(6). In a study of fresh produce packing sheds, Ailes found that produce samples taken from

packing shed bins, boxes, turntables, and conveyor belts had significantly greater likelihood of

E. coli contamination than had those taken from the field (1). A survey of non-egg-contact

surfaces observed that surfaces such as walls, floors, drains, air handling apparatus, and forklifts

can harbor organisms that can be transferred to egg-contact surfaces and eggs themselves during

production and between cleaning and sanitation periods (56). In a 1999 outbreak of Salmonella

15

Agona, investigators found widespread low levels of the organism in the plant environment,

including samples taken from the floor, production equipment, and the exhaust system in the

cereal plant implicated (58).

Attachment of bacteria to surfaces can be categorized as short-term attached or dried on

(approximately l-2 hours), mid-term attached (approximately the length of a production shift e.g.

several hours) and long term attached which is often termed biofilm growth (30). Long term

bacterial attachment on equipment surfaces enhances the potential of the surfaces as a source of

food contamination.

The survivability of non-biofilm Salmonella on a surface was found to be dependent

upon relative humidity, soil level, type of soil and type of surface (29). Allan showed that

Salmonella can survive on stainless steel, fiberglass reinforced plastic (FRP), and acetal resin for

up to 15 days at 10°C and high relative humidity (2). Kusumaningrum et al. showed Salmonella

Enteritidis was able to be recovered from dry surfaces for at least 4 days at high (105 CFU/cm

2)

contamination levels, but at moderate level (103 CFU/cm

2), the numbers decreased to the

detection limit within 24 h and at low level (10 CFU/cm2) within 1 h (39). D-values of 1363.2,

481.8, and 134.2 min were calculated for Salmonella cells suspended in TSB and then spotted on

formica, stainless steel, and ceramic tile surfaces, respectively (12).

Surfaces from which it is easier to remove food debris and microorganisms are generally

considered the most hygienic and safe. However, a study by Moore et al. demonstrated that the

characteristics that enable a surface to be easily cleaned may also render it more likely to release

organisms to food (54). Out of nine food contact surfaces tested, stainless steel and PVC

required the most time for Pseudomonas cells to adhere and double (7). Moore et al. showed

that Salmonella inoculated and dried onto stainless steel transferred at rates of 36 to 66% and 23

16

to 31% for wet and dry Romaine lettuce, respectively (53). S. aureus inoculated at 7 log

CFU/cm2 onto chicken meat was found to transfer to stainless steel and polyethylene at

populations of 4.25 and 4.46, respectively in 10 s contact time (16). S. enteritidis exhibited

instantaneous transfer rates of 94±42% and 105±26% to roasted chicken filets and cucumber

slices respectively when 500 g/slice force was applied (39). Salmonella transfer can also occur

from equipment onto edible plant tissues during slicing or into the juice of freshly pressed fruits

(20).

17

References

1. Ailes, E. C., J. S. Leon, L. A. Jaykus, L. M. Johnston, H. A. Clayton, S. Blanding, D. G.

Kleinbaum, L. C. Backer, and C. L. Moe. 2008. Microbial concentrations on fresh

produce are affected by postharvest processing, importation, and season. J. Food Prot.

71:2389-2397.

2. Allan, J. T., Z. Yan, and J. L. Kornacki. 2004. Surface material, temperature, and soil

effects on the survival of selected foodborne pathogens in the presence of condensate. J.

Food Prot. 67:2666-2670.

3. Behling, R. G., J. Eifert, M. C. Erickson, J. B. Gurtler, J. L. Kornacki, E. Line, R.

Radcliff, E. T. Ryser, B. Stawick, and Z. Yan. 2010. Selected pathogens of concern to

industrial food processors: infectious, toxigenic,toxico-infectious, selected emerging

pathogenic bacteria. In J.L. Kornacki (ed.), Principles of microbiological troubleshooting

in the industrial food processing environment. Springer Science+Business Media, LLC,

New York, NY.

4. Block, S. S. (ed.). 2001. Disinfection, sterilization, and preservation. Lippincott

Williams & Wilkins, Philadelphia, PA.

5. Boulange Petermann, L. 1996. Processes of bioadhesion on stainless steel surfaces and

cleanability: a review with special reference to the food industry. Biofouling. 10:275-

300.

6. Brackett, R. E. 1999. Incidence, contributing factors, and control of bacterial pathogens

in produce. Postharvest Biol. Technol. 15:305-311.

7. Careli, R. T., N. J. Andrade, N. F. Soares, J. I. Ribeiro, M. S. Rosado, and P. C.

Bernardes. 2009. The adherence of Pseudomonas fluorescens to marble, granite,

synthetic polymers, and stainless steel. Cienc. Tecnol. Aliment. 29:171-176.

8. Casani, S., M. Rouhany, and S. Knochel. 2005. A discussion paper on challenges and

limitations to water reuse and hygiene in the food industry. Water Res. 39:1134-1146.

9. Centers for Disease Control and Prevention. 2010. Preliminary FoodNet data on the

incidence of infection with pathogens transmitted commonly through food- 10 states,

2009. In, Morbidity and Mortality Weekly Report, vol. 59.

10. Centers for Disease Control and Prevention. 2010. Salmonella homepage. Available at:

http://www.cdc.gov/salmonella/general/index.html. Accessed 14 October 2010.

11. Centers for Disease Control and Prevention. 2010. Salmonella outbreaks. Available at:

http://www.cdc.gov/salmonella/outbreaks.html. Accessed 14 October 2010.

http://www.cdc.gov/salmonella/general/index.html

http://www.cdc.gov/salmonella/outbreaks.html

18

12. Cesare, A. d., B. W. Sheldon, K. S. Smith, and L.-A. Jaykus. 2003. Survival and

persistence of Campylobacter and Salmonella species under various organic loads on

food contact surfaces. J. Food Prot. 66:1587-1594.

13. Chaim, A., M. Pessoa, and V. L. Ferracini. 2002. Pesticide efficiency deposition

obtained with electrostatic spray head for knapsack mistblower sprayer. Pesqu.

Agropecu. Bras. 37:497-501.

14. Cooper, S. C., and S. E. Law. 2006. Electrostatic sprays for sunless tanning of the

human body. IEEE Trans. 42:385-391.

15. Cramer, M. M. 2006. Food plant sanitation design, maintenance, and good

manufacturing practices. CRC Press, Boca Raton, FL.

16. da Silva Malheiros, P., C. T. dos Passos, L. S. Casarin, L. Serraglio, and E. C. Tondo.

2010. Evaluation of growth and transfer of Staphylococcus aureus from poultry meat to

surfaces of stainless steel and polyethylene and their disinfection. Food Control.

21:298-301.

17. Dai, Y., S. C. Cooper, and S. E. Law. 1992. Effectiveness of electrostatic, aerodynamic

and hydraulic spraying methods for depositing pesticide sprays onto inner plant regions

and leaf undersides. Trans. ASAE. No. 92-1094, Charlotte, NC.

18. Davies, J., C. S. Nunnerley, A. C. Brisley, J. C. Edwards, and S. D. Finlayson. 1996.

Use of dynamic contact angle profile analysis in studying the kinetics of protein removal

from steel, glass, polytetrafluoroethylene, polypropylene, ethylenepropylene rubber, and

silicone surfaces. J. Colloid Interface Sci. 182:437-443.

19. Detry, J. G., M. Sindic, and C. Deroanne. 2010. Hygiene and cleanability: a focus on

surfaces. Crit. Rev. Food Sci. Nutr. 50:583-604.

20. Doyle, M. P., and L. R. Beuchat (ed.). 2007. Food microbiology: fundamentals and

frontiers. ASM Press, Washington, D.C.

21. Durr, H. 2007. Influence of surface roughness and wettability of stainless steel on soil

adhesion, cleanability and microbial inactivation. Food Bioprod. Process. 85:49-56.

22. Environmental Protection Agency. 1976. Sanitizer test for inanimate surfaces.

Available at: http://www.epa.gov/oppad001/dis_tss_docs/dis-10.htm. Accessed 14

October 2010.

23. Environmental Protection Agency. 1979. Efficacy data requirements sanitizing rinses

(for previously cleaned food-contact surfaces). Available at:

http://www.epa.gov/oppad001/dis_tss_docs/dis-04.htm. Accessed 14 October 2010.

http://www.epa.gov/oppad001/dis_tss_docs/dis-10.htm

http://www.epa.gov/oppad001/dis_tss_docs/dis-04.htm.

19

24. Evans, M. D., S. E. Law, and S. C. Cooper. 1994. Fluorescent spray deposit

measurement via light intensified machine vision. Trans. ASAE. 10:441-447.

25. Food and Drug Administration. 2010. Code of Federal Regulations, 21CFR110.3.

Available at: http://www.fsis.usda.gov/OPPDE/rdad/FRPubs/00-014R/fda-gmpregs.htm.

Accessed 14 October 2010.

26. Frost, A. R., and S. E. Law. 1981. Extended flow characteristics of the embedded-

electrode spray-charging nozzle. J. Agric. Engineer. Research. 26:79-86.

27. Gibson, H., R. Elton, W. Peters, and J. T. Holah. 1995. Surface and suspension testing:

Conflict or complementary. Int. Biodeterior. Biodegrad. 36:375-384.

28. Giles, D. K. and S.E. Law. 1985. Space charge deposition of pesticide sprays onto

cylindrical target arrays. Trans. ASAE. 28:658-664.

29. Helke, D. M., and A. C. L. Wong. 1994. Survival and growth characteristics of Listeria

monocytogenes and Salmonella typhimurium on stainless steel and buna-N rubber. J.

Food Prot. 57:963-968.

30. Holah, J. T., A. Lavaud, W. Peters, and K. A. Dye. 1998. Future techniques for

disinfectant efficacy testing. Int. Biodeterior. Biodegrad. 41:273-279.

31. Holah, J. T., J. H. Taylor, D. J. Dawson, and K. E. Hall. 2002. Biocide use in the food

industry and the disinfectant resistance of persistent strains of Listeria monocytogenes

and Escherichia coli. J. Appl. Microbiol. 92:111S-120S.

32. Hsu, S.-Y., C. Kim, Y.-C. Hung, and S. E. Prussia. 2004. Effect of spraying on chemical

properties and bactericidal efficacy of electrolysed oxidizing water. Int. J. Food Sci.

Technol. 39:157-165.

33. Hui, Y. H., B. L. Bruinsma, J. R. Gorham, W.-K. Nip, P. S. Tong, and P. Ventresca.

2003. Food plant sanitation. Marcel Dekker, Inc., New York, NY.

34. Kang, T.-G., D.-H. Lee, C.-S. Lee, S.-H. Kim, G.-I. Lee, W.-K. Choi, and S.-Y. No.

2004. Spray and depositional characteristics of electrostatic nozzles for orchard sprayers.

ASAE Annual Meeting. Paper number 041005.

35. Katsuyama, A. M., and J. P. Strachan (ed.). 1980. Principles of food processing

sanitation. The Food Processors Institute, Washington, D.C.

36. Kim, C., and Y. C. Hung. 2007. Development of a response surface model of an

electrostatic spray system and its contributing parameters. Trans. ASABE. 50:583-590.

http://www.fsis.usda.gov/OPPDE/rdad/FRPubs/00-014R/fda-gmpregs.htm

20

37. Kim, H., J.-H. Ryu, and L. R. Beuchat. 2007. Effectiveness of disinfectants in killing

Enterobacter sakazakii in suspension, dried on the surface of stainless steel, and in a

biofilm. Appl. Environ. Microbiol. 73:1256-1265

38. Knight, G. C., and H. M. Craven. 2010. A model system for evaluating surface

disinfection in dairy factory environments. Int. J. Food Microbiol. 137:161-167.

39. Kusumaningrum, H. D., G. Riboldi, W. C. Hazeleger, and R. R. Beumer. 2003.

Survival of foodborne pathogens on stainless steel surfaces and cross-contamination to

foods. Int. J. Food Microbiol. 85:227-236.

40. Latheef, M. A., J. B. Carlton, I. W. Kirk, and W. C. Hoffmann. 2009. Aerial

electrostatic-charged sprays for deposition and efficacy against sweet potato whitefly

(Bemisia tabaci) on cotton. Pest Manag. Sci. 65:744-752.

41. Law, S. E. 1978. Embedded-electrode electrostatic-induction spray-charging nozzle -

theoretical and engineering design. Trans. ASAE. 21:1096-1104.

42. Law, S. E. 1983. Electrostatic pesticide spraying: concepts and practice. IEEE Trans.

Ind. Appl. IA-19:160-168.

43. Law, S. E., and S. C. Cooper. 2001. Air-assisted electrostatic sprays for postharvest

control of fruit and vegetable spoilage microorganisms. IEEE Trans. 37:1597-1602.

44. Law, S. E., S. C. Cooper, and M. A. Harrison. 2004. Electrostatic spray application of

decontaminant agents onto the human body as a bioterrorism countermeasure: process

development and evaluation. p. 331-336. In H. Morgan (ed.), Electrostatics 2003:

proceedings of the Electrostatics Conference of the Institute of Physics: held in

Edinburgh, UK, 23-27 March, 2003. CRC Press LLC, Boca Raton, FL.

45. Law, S. E., and M. D. Lane. 1981. Electrostatic deposition of pesticide spray onto foliar

targets of varying morphology. Trans. ASAE. 24:1441-1445, 1448.

46. Law, S. E., H. Y. Wetzstein, S. Banerjee, and D. Eisikowitch. 2000. Electrostatic

application of pollen sprays: effects of charging field intensity and aerodynamic shear

upon deposition and germinability. IEEE Trans. 36:998-1009.

47. Lelieveld, H. L. M., M. A. Mostert, J. Holah, and B. White (ed.). 2000. Hygiene in food

processing. CRC Press LLC, Boca Raton, FL.

48. Marriot, N. G., and R. B. Gravani. 2006. Principles of food sanitation. Springer

Science+Business Media, Inc., New York, New York.

49. Maski, D., and D. Durairaj. 2010. Effects of charging voltage, application speed, target

height, and orientation upon charged spray deposition on leaf abaxial and adaxial

surfaces. Crop Prot. 29:134-141.

21

50. Maski, D., and D. Durairaj. 2010. Effects of electrode voltage, liquid flow rate, and

liquid properties on spray chargeability of an air-assisted electrostatic-induction spray-

charging system. J. Electrostatics. 68:152-158.

51. Matthews, G. A. 1989. Electrostatic spraying of pesticides - a review. Crop Prot. 8:3-

15.

52. Midelet, G., and B. Carpentier. 2002. Transfer of microorganisms, including Listeria

monocytogenes, from various materials to beef. Appl. Environ. Microbiol. 68:4015-

4024.

53. Moore, C. M., B. W. Sheldon, and L. A. Jaykus. 2003. Transfer of Salmonella and

Campylobacter from stainless steel to romaine lettuce. J. Food Prot. 66:2231-2236.

54. Moore, G., I. S. Blair, and D. A. McDowell. 2007. Recovery and transfer of Salmonella

typhimurium from four different domestic food contact surfaces. J. Food Prot. 70:2273-

2280.

55. Moretro, T., E. S. Midtgaard, L. L. Nesse, and S. Langsrud. 2003. Susceptibility of

Salmonella isolated from fish feed factories to disinfectants and air-drying at surfaces.

Vet. Microbiol. 94:207-217.

56. Musgrove, M. T., D. R. Jones, J. K. Northcutt, P. A. Curtis, K. E. Anderson, D. L.

Fletcher, and N. A. Cox. 2004. Sources and risk factors for contamination, survival,

persistence, and heat resistance of Salmonella in low-moisture foods. J. Food Prot.

67:1919-1936.

57. Pérez-Rodríguez, F., A. Valero, E. Carrasco, R. M. García, and G. Zurera. 2008.

Understanding and modelling bacterial transfer to foods: a review. Trends Food Sci.

Technol. 19:131-144.

58. Podolak, R., E. Enache, W. Stone, D. G. Black, and P. H. Elliot. 2004. Survey of shell

egg processing plant sanitation programs: effects on non-egg-contact surfaces. J. Food

Prot. 67:2801-2804.

59. Polat, M., H. Polat, and S. Chander. 2000. Electrostatic charge on spray droplets of

aqueous surfactant solutions. J. Aerosol Sci. 31:551-562.

60. Pordesimo, L. O., E.G. Wilkerson, A.R. Womac, and C.N. Cutter. 2002. Process

engineering variables in the spray washing of meat and produce. J. Food Prot. 65:222-

237.

61. Queensland Australia Department of Employment, Economic Development and

Innovation and Department of Environment and Resource Management Production.

2010. Water efficiency overview - W1. Available at:

22

http://www.ecoefficiency.com.au/Portals/56/factsheets/foodprocess/water/ecofoodwater_

fsw1.pdf. Accessed 14 October 2010.

62. Russell, S. 2003. The effect of electrolyzed oxidative water applied using electrostatic

spraying on pathogenic and indicator bacteria on the surface of eggs. Poult. Sci. 82:158-

162.

63. Sansebastiano, G., R. Zoni, and L. Bigliardi. 2007. Cleaning and disinfection procedures

in the food industry general aspects and practical applications. p. 253-280. In A.

McElhatton, and R.J. Marshall (ed.), Food safety: a practical and case study approach.

Springer Science+Business Media, LLC, New York, NY.

64. Scallan E, Griffin PM, Angulo FJ, Tauxe RV, Hoekstra RM. January 2011. Foodborne

illness acquired in the United States—unspecified agents. Emerg Infect Dis [serial on the

internet]. Available at: http://www.cdc.gov/EID/content/17/1/16.htm. Accessed 31

March 2011.

65. Scallan E, Hoekstra RM, Angulo FJ, Tauxe RV, Widdowson M-A, Roy SL, et al.

January 2011. Foodborne illness acquired in the United States—major pathogens.

Emerg Infect Dis [serial on the internet]. Available at:

http://www.cdc.gov/EID/content/17/1/7.htm. Accessed 31 March 2011.

66. Scherm, H., A. T. Savelle, and S. E. Law. 2007. Effect of electrostatic spray parameters

on the viability of two bacterial biocontrol agents and their deposition on blueberry

flower stigmas. Biocontrol Sci. and Technol. 17:285-293.

67. Starrs, C. 2009. New applications help company with patented spray system grow.

Available at: http://www.onlineathens.com/stories/030109/bus_399637493.shtml.

Accessed 14 October 2010.

68. United States Department of Agriculture. 2010. Foodborne illness cost calculator:

Salmonella. Available at:

http://www.ers.usda.gov/Data/FoodborneIllness/salm_Intro.asp. Accessed 14 October

2010.

69. van Klingeren, B., W. Koller, S. F. Bloomfield, R. Böhm, A. Cremieux, J. Holah, G.

Reybrouck, and H. J. Rödger. 1998. Assessment of the efficacy of disinfectants on

surfaces. Int. Biodeterior. Biodegrad. 41:289-296.

70. Washington Metropolitan Area Transit Authority. 2009. ESS - D.C. Metro prepares for

Swine Flu. Available at: http://www.youtube.com/watch?v=l3kmRQx8m3A. Accessed

14 October 2010.

http://www.ecoefficiency.com.au/Portals/56/factsheets/foodprocess/water/ecofoodwater_fsw1.pdf

http://www.ecoefficiency.com.au/Portals/56/factsheets/foodprocess/water/ecofoodwater_fsw1.pdf

http://www.cdc.gov/EID/content/17/1/16.htm


http://www.onlineathens.com/stories/030109/bus_399637493.shtml

http://www.ers.usda.gov/Data/FoodborneIllness/salm_Intro.asp

http://www.youtube.com/watch?v=l3kmRQx8m3A.

23

Chapter 3


METHOD FOR FOOD PROCESSING SURFACES1

______________________________________

1Shawn M. Lyons, Mark A. Harrison, and S. Edward Law. To be submitted to Journal of Food

Protection

24

Abstract

In this study, air-assisted, induction-charged sprays of sanitizers were applied to

inoculated food contact surfaces to evaluate their ability to reduce populations of Salmonella.

Electrostatically charged sprays (-7.2 mC/kg charge-to-mass ratio) deposited more active

ingredient and carrier liquid onto target surfaces than uncharged sprays from the same nozzle and

a conventional hydraulic nozzle (p<0.05). Charged sprays at lowered biocide rates reduced

Salmonella population on target surfaces greater than or equal to hydraulic sprays with full-rate

biocide for 8 of 9 surfaces and orientations combinations evaluated (p<0.05). Peracetic acid

sprays were more effective than quaternary ammonium compound sprays from all nozzles in 9 of

9 surface and orientation combinations evaluated (p<0.05).

Index words: electrostatic spray, quaternary ammonium compounds, peracetic acid, food contact

surfaces, Salmonella.

25

Introduction and Literature Review

Foodborne pathogens are a significant contributor to disease in the United States, causing

food borne illness and economic loss. Scallan et al. estimated 38.4 million episodes of

domestically acquired foodborne illness were caused by unspecified agents, resulting in 71,878

hospitalizations and 1,686 deaths (38). Additionally, Scallan et al. identified 31 major pathogens

which were responsible for and estimated 9.4 million episodes of foodborne illness 55,961

hospitalizations, and 1,351 deaths. Of these incidences caused by specific agents, nontyphoidal

Salmonella spp. were the leading cause of bacterial foodborne illness, 11% of total. Among all

pathogens, nontyphoidal Salmonella spp. were the leading cause of hospitalizations and deaths,

35 and 28% respectively (39). In 2009 USDA‘s Economic Research Service (ERS) estimated

the number of Salmonella cases due to any source as 1.4 million with an associated cost of 2.6

billion USD per year (42). According to the Centers for Disease Control and Prevention, in 2009

a total of 17,468 laboratory-confirmed cases of foodborne pathogen infection were identified.

Salmonella species accounted for 7,039 reported infections, having an incidence of 15.19 per

100,000 population (6).

Salmonella contamination in the food factory environment is due to their ubiquitous

presence in the natural environment and likely presence in some raw ingredients (3). In the past

three years, multi-state outbreaks of foodborne Salmonella infection have been associated with

food items such as: alfalfa sprouts, cheesy chicken rice frozen entrée, red and black pepper,

Italian style meats, pistachios, cantaloupes, rice and wheat cereals, peanut butter, and raw

produce (7). Presence of pathogenic bacteria in foods increases the probability of those bacteria

transferring to food contact surfaces and the cross contamination of other food items.

26

Once in a food factory environment, the number of Salmonella can multiply based on the

conditions and control measures in place (5). In a study of fresh produce packing sheds, Ailes

found that produce samples taken from the bin, box, turntable, and conveyor belt packing shed

steps had significantly greater likelihood of E. coli contamination than had those taken from the

field (1).

A survey of non-egg-contact surfaces observed that surfaces such as walls, floors, drains,

air handling apparatus, and forklifts can harbor organisms that can be transferred to egg-contact

surfaces and eggs themselves during production and between cleaning and sanitation periods

(33). In a 1999 outbreak of Salmonella Agona, investigators found widespread low levels of the

organism in the plant environment, including samples taken from the floor, production

equipment, and the exhaust system in the cereal plant implicated (32).

Attachment of bacteria to surfaces can be categorized as short-term attached or dried on

(approximately l-2 hours), mid-term attached (approximately the length of a production shift e.g.

several hours) and long term attached which is often termed biofilm growth (16). The

survivability of nonbiofilm Salmonella on a surface was found to be dependent upon relative

humidity, soil level, type of soil and type of surface (15).

Cleaning and sanitizing food processing surfaces can be divided into several stages:

wetting and penetration of the surfaces with water and cleaning solution, the reaction of the

cleaning solution with the soil and surface, the prevention of redeposition back onto the cleansed

surface, the wetting by the sanitizing solution of residual microorganisms to produce a biocidal

or biostatic action, and dispersion of microorganisms if needed (27). The most common cleaning

procedures include manual application, immersion/clean-out-of-place (COP), and clean-in-place

(CIP) (37).

27

There are many methods by which biocides suspended in water can be applied to a

surface. These methods include spray sanitizing, fogging, and flood sanitizing. Spray or mist

sanitizing and flood sanitizing are applied via spray nozzles, but differ in the fact that flood

sanitizing methods apply large quantities of sanitizer to ensure extensive exposure which

increases costs of sanitizer and water while simultaneously creating a wet condition (28). Spray

or mist sanitizing is the most commonly used method for applying sanitizers. The efficiency of

conventional spraying depends on (i) the water pressure employed, (ii) the volume of water used,

(iii) the water temperature, (iv) the distance of the target from the spray origin (contact force), (v)

the time of exposure of the target to the spray, (vi) the type and number of spray nozzles used,

(vii) the nozzle orientation, and (viii) the spray pattern (36). In the same survey 52% of food

factories used a conventional hand sprayer or other type of sprayer and 28% used high pressure

sprayers including foamers out of 117 sites (17).

Law developed an embedded-electrode electrostatic-induction spray-charging nozzle to

enhance the deposition of liquid pesticide droplets having a low to middle range resistivity of 10-

1 to 10

4 ohm m onto agricultural plants (22). The addition of a pneumatic force of as low as 30

psi shears this liquid into a film which atomizes into negatively charged droplets and then propel

the spray liquid toward the desired target (22). The aforementioned nozzle initially yielded an

average droplet charge-to-mass ratio of 4.8 mC/kg and later design improvements tripled this

value. Factors that affect electrostatic spray deposition include: charge-to-mass ratio, applied

voltage, atomizing air pressure, liquid flow rates, and liquid resistivity (13, 22). An increased

charge-to-mass ratio results in increased deposition and ―wrap-around‖ effect. Fluorometric

analysis of tracer spray deposits on target surfaces has been a common method of measuring

performance of sprayers on variety of surface types and morphologies (12).

28

Microbiological assessment in the same air-assisted induction-charged electrostatic

spraying system showed a charged spray of quaternary ammonium onto Pseudomonas inoculated

targets placed around human body sites to have an average 2.38 log CFU reduction compared to

uncharged sprays with 0.66 log CFU reduction (24). Hsu et al. applied electrolyzed water via

this electrostatic spray gun to produce a 3-4 log CFU reduction of Listeria monocytogenes on

surfaces (18). Russell similarly applied electrolyzed water to unhatched eggs inoculated with

Salmonella to produce at least a 4 log CFU reduction over the control treatment (36). In recent

years, this type of hand-held electrostatic spraying equipment has been used in efforts to

disinfect schools, hospitals, hotels (40) and transit systems (42).

The objective of this study was to evaluate the mass transfer efficiency and the

antimicrobial efficacy of biocides applied as an air-assisted, induction-charged spray. For two

sanitizers, an equivalence point was determined at which a reduced mass of active ingredient

dispensed from an air-assisted electrostatic nozzle with charging on has the same effect in terms

of population reduction of Salmonella as a conventional hydraulic-atomizing spray nozzle at full

rate mass of active ingredient. Using the equivalence point mass of sanitizer dispensed toward

target coupons at a reduced volume of carrier liquid, efficacy of nozzle treatments was evaluated

in terms of population reduction of Salmonella using two liquid sanitizers, applied to three food

contact surface materials, in three target orientations.

Materials and Methods

Spray Delivery Systems

The electrostatic charging pneumatic atomizing spray nozzle used in the study was

developed by Law (22) and provided by Electrostatic Spraying Systems, Inc. (Watkinsville, GA).

The nozzle uses induction charging and pneumatic energy to produce finely atomized charged

29

droplets with a volume median diameter of 30 micrometers. Compressed air at 30 psig from

laboratory source was used to atomize droplets. Atomizing air pressure was valve controlled and

monitored using a 0-60 psi pressure gauge on the air line. A Lambda model LLS6108 0-18 Vdc

(Lambda, San Diego, CA) was used as a power source and amplified by a Venus high voltage

power supply (0-1500 Vdc output) to provide a positive 1200 Vdc supply to the embedded

induction electrode of the spray nozzle for spray charging. The resultant spray cloud carried a

negative current which was measured in -μA using a digital multimeter (Extech model 410,

Waltham, MA). The cloud current was measured using a 26 gauge needle ionization probe held

2.54 cm. from nozzle in center of spray cloud. The average charge-to-mass ratio of the spray

cloud was calculated using current on spray cloud and liquid flow rate. A charge-to-mass ratio

of -7.2 mC/kg was imparted to the air-assisted, inductively-charged sprays. No induction voltage

(0 mC/kg) was imparted by the electrostatic nozzle with charging off treatments. Spray liquids

were contained in a 500 mL reservoir which sat on a stirrer to keep spray liquids mixed.

The hydraulic-atomizing nozzle used in the study was a Spraying Systems Co.

(Springfield, IL) Teejet Even Flat Spray Tip TP40015E with a slotted strainer part 451-NY-20

producing spray having median size of 300 µm. The spray tip dispensed 600 mL/min at 43 psi

hydraulic pressure monitored by a gauge (Weksler, Deer Park, NY ). Spray liquid was contained

in an 8 liter reservoir (Nalgene, Rochester, NY) with an on/off valve. Hydraulic pressure was

provided by a twin piston pump attached to a motor (Dayton model 5K121AF, USA). The flat

spray tip selected produced a 30.48 x 10.16 cm vertical spray pattern at a nozzle to target

distance of 42 cm.

30

Spraying Apparatus and Target Array

The spraying area was in a biosafety level 2 laboratory, using a modified stainless steel

smokehouse oven (Alkar model 450,Alkar-RapidPak Inc., Lodi, WI) as a spray chamber (1.5 m

high x 1 m wide x 1 m deep). The door of the chamber was removed to create a large

containment area with an exhaust fan. A plastic dam 2.54 cm in height was sealed onto the floor

in the front face of the chamber in order to contain liquids within the oven. Two large acrylic

facings were fabricated in order to cover unneeded area on the face of the chamber thus

increasing exhaust velocity at its entry.

A plywood rack was constructed and coated in water-proof paint to hold spraying

equipment and house a robotic arm which provided mechanical repeatability of spray treatments.

The robotic arm consisted of a 0.635 cm. diameter stainless steel tube 99 cm in length centered

on the spray rack. The arm created an arc shaped spray path of 120° which passed through the

center of each spray target coupon. The robotic arm moved from a stationary position left-to-

right and right-to-left in a dual pass for all treatments. Each dual pass was electronically

controlled to occur in a total of 6000±200 ms. PVC piping was attached to the side of the

chamber to collect spray and direct it downward into the chamber‘s floor while the robotic arm

was stationary.

A target coupon holding arc was affixed to a horizontal sliding mechanism which

allowed for rapid adjustment of target spacing for the electrostatic and hydraulic nozzle,

respectively. The target holding arc contained four equidistant holes spaced 15.24 cm from one

another and thumb screws for placement and securing of target coupons which were placed in

the target arc using metal alligator clips. Target coupons were evaluated in two different

positions: parallel and perpendicular to incoming spray vector. The metal target arc and coupons

31

affixed to the arc were grounded earth using a wire, ensuring metal to metal contact throughout

the system.

Aerosol contamination of laboratory surfaces was contained by running the chamber‘s

exhaust fan at full capacity during spray treatments producing approximately 0.28 m3/min of air

flow. Pathogen contamination of inside chamber surfaces was contained and eliminated by post-

experiment flood sanitization using quaternary ammonium compound Zep FS Formula 386L

(Zep, Atlanta, GA).

Experiment 1: Fluorometric Mass Transfer Analysis

A fluorometer (Turner model 450, Sunnyvale, CA) was used to quantify mass of blaze

orange tracer (DayGlo Color Corp., Cleveland, OH) transferred from spray nozzle to targets on a

scale of 0-2000 units. The fluorometer was calibrated using known concentrations of blaze

orange tracer.

In order to quantify mass of blaze orange tracer deposited onto target coupons given the

apparatus built for the experiment, standard tracer solutions were prepared, sprayed, reclaimed,

and read fluorometrically. Clean stainless steel target coupons were used as idealized targets for

mass transfer experiments. Mass transfer was done for two orientations, perpendicular to nozzle

spray vector and parallel to spray vector. Perpendicular and parallel to spray vector orientations

contain two sides in which tracer suspension was deposited, front side/back side and left

side/right side, respectively. Left side and right side were treated statistically as the same

surface. In each orientation, the two target coupons were clipped together using metal alligator

clips and edges sealed together using Teflon pipe tape. Teflon tape prevented tracer suspension

from seeping between the target coupons and giving falsely increased results. Teflon tape

covered 15% of the target coupon surface.

32

Standard tracer solutions were sprayed from the corresponding three nozzles under

standard spraying conditions. Per each spray nozzle type, pairs of target coupons were clipped

together and placed in four arc positions for a total of 8 samples per spray nozzle dual pass

treatment. After spraying, the Teflon tape was carefully removed with forceps and the stainless

steel targets were placed in separate 80 ml wash vessels (Nalgene, Rochester, NY). Thirty ml of

wash solution was added to each vessel to reclaim tracer from the target coupons and suspend it

into liquid for reading in the fluorometer. Wash vessels were tumbled for ten minutes.

Recovered tracer suspension was transferred into clean 12 x 75 mm glass cuvettes (VWR, West

Chester, PA) using clean 14.6 cm glass pipets (Fisher, USA). Three fluorometer readings were

taken from each wash vessel, dumping the tracer suspension from the cuvette between readings.

Fluorometer readings over 2000 were diluted in a 1:1 tracer suspension to wash solution ratio

and multiplied resultant readings by a factor of two. Once the concentration of tracer deposited

in μg/liter was calculated from the calibration curve equation, the mass of blaze orange tracer in

μg was calculated by multiplying by 0.03, or the volume of wash liquid used to reclaim the tracer

from target coupon surfaces.

Preparation of Tracer Suspension and Wash Solution

The fluorescent industrial pigment, blaze orange was used in the study as a tracer in order

to quantify the mass transferred to target coupons for each spray nozzle type through

fluorometric analysis. Blaze orange is an insoluble, inert, powder which is 4-5 µm in size and

has no effect on suspension electric conductivity. A surfactant, Triton® X-100 (Fisher

Scientific, USA) was used to suspend the tracer powder in deionized water (DI water). To

standardize conductivity, 0.1 g/liter sodium chloride was added to solutions. The spray solutions

were prepared as follows:

33

Electrostatic tracer suspension: 4.5 g Blaze orange tracer and 1.5 ml Triton X-100

surfactant into 1,500 ml deionized water.

Hydraulic tracer suspension: 500 ml electrostatic tracer suspension into 2,500 ml

deionized water.

Sodium chloride (0.1 g/liter) was added to each suspension after blaze orange tracer was

adequately suspended in DI water. Target wash solution was prepared by adding 1.0 ml of

Triton® X-100 to 1 liter DI water.

Experiment 2: Microbiological equivalence point

For both peracetic acid and quaternary ammonium sanitizers, the rate of biocide at which

bacterial population reduction on target coupons sprayed using an electrostatic nozzle was

equivalent to the bacterial population reduction of target coupons sprayed at full rate hydraulic-

atomizing nozzle was determined. An equivalence curve was constructed which plotted

Salmonella population log reduction values of full rate hydraulic-atomizing nozzle and

electrostatic nozzle rates of 0, 1/8, 1/6, 1/4, 1/2, and 1 (full rate). Full rate hydraulic-atomizing

nozzle sanitizer was prepared according to manufacturers‘ instructions for food contact surfaces,

0.26-0.28% v/v concentration and 200 ppm for peracetic acid and quaternary ammonium

compound, respectively. Quaternary ammonium compound sanitizer levels were tested with a

titration kit (Quat test kit #317, Ecolab, St. Paul, MN) to verify amount of active ingredient. Full

rate electrostatic nozzle sanitizer was prepared with six times the active ingredient of hydraulic-

atomizing sanitizer to correct for the discrepancy in spray volumes dispensed (100 ml/min

electrostatic vs. 600 ml/min hydraulic) thus ensuring the same mass of active ingredient was

dispensed toward the target coupon face. Dilutions of full rate electrostatic nozzle sanitizer were

made using sterile tap water.

34

Sterile tap water was used as a control spray treatment to quantify the amount of

Salmonella removed from target coupon faces due to physical ―wash off‖. Stainless steel target

coupons were inoculated to with Salmonella to a level of 107 CFU/coupon face. For equivalence

study, front side stainless steel target coupons were used as idealized targets. In each spray

treatment, four target coupons pairs were placed in the arc holder and sprayed under standard

operating conditions. Only front side coupons were inoculated with Salmonella. All treatments

were replicated three times for each sanitizer.

Experiment 3: Application of Equivalence Rate to Various Surfaces and Orientations

Additional food contact surface types and target coupon orientations inoculated with

Salmonella were challenged using the equivalence rate found for sanitizers in experiment 2.

Food contact surfaces stainless steel, polyvinyl chloride #120, and waxed cardboard were

evaluated in two orientations: perpendicular and parallel to incoming spray vector.

Perpendicular orientation contained two inoculated faces, front side and back side. Parallel

orientation contained two inoculated faces, left side and right side. Sanitizers for use in

electrostatic nozzle treatments were prepared at calculated equivalence rates in sterile tap water.

Sanitizers for use in hydraulic-atomizing nozzle were used at full rate. Sterile tap water was

used as a control spray treatment to quantify the amount of Salmonella removed from target

coupon faces due to physical ―wash off‖. In each spray treatment, eight target coupons were

placed in four arc holder positions, two sides per position, and sprayed under standard operating

conditions. All treatments were replicated three times for each spray liquid, sanitizer, and tap

water control.

35

Salmonella enterica Serovars

Salmonella enteric serovars Baildon, Enteritidis, Poona, St. Paul, and Typhimurium were

obtained from the culture collection in the Department of Food Science and Technology,

University of Georgia, Athens. Cultures from frozen storage were activated by three consecutive

loopful transfers into 9 ml tryptic soy broth (Becton Dickinson and Company, Sparks, MD) at

24 h intervals and incubated at 37°C. Prior to spray treatments, a loopful of each serovar was

transferred and incubated at 37°C for 18-24 h. On the day of the experiment, prior to inoculation

of target coupons, 2 ml of each serovar was pooled into a cocktail. The 10 ml cocktail was

centrifuged (Beckman Coulter Allegra X-22R, Fulleton, CA) at a relative centrifugal force

(RCF) of 2300 x g for 5 min at 22°C and the supernatant removed. The pellet was resuspended

via vortex in 10 ml of 0.1% peptone water (Becton Dickinson) and centrifuged for another 5 min

at 2300 x g and 22°C, removing the supernatant once again. The pellet was resuspended via

vortex in 10 ml of 0.1% peptone water (Becton Dickinson) for final use. This resuspended

culture was used as a stock culture to make 1:10 dilutions in 0.1% peptone water for inoculation

if target coupons. Resuspended cultures contained approximately 108 CFU/ml Salmonella.

Target Coupon Fabrication and Inoculation

Target coupons sized 23x90 mm were fabricated from stainless steel, polyvinyl chloride

(PVC) belting, and waxed cardboard. Stainless steel coupons type 304, finish #4B, thickness

0.9 mm were sheared to size from sheets by University of Georgia Instrument Shop, Athens GA

and the edges smoothed. They were degreased using acetone (Sigma-Aldrich, St. Louis, MO)

when acquired. The coupons were cleaned by placing them in 100 ml/liter solution of Micro-90

(International Products Corporation, Burlington, NJ) at 80°C for 1 h in an ultrasonic bath (VWR

model 550HT, USA). Coupons were then rinsed in deionized water and immersed in 1.5%

36

phosphoric acid solution (Fisher Scientific, USA) at 80°C for 20 min. Coupons were then rinsed

in deionized water, and immersed in deionized water for autoclave sterilization. Polyvinyl

chloride belting (#120 white; W.L. Deckert Co., Inc., Milwaukee, WI) was cut to size, 24 x 90

cm, 3 mm thick using scissors. All cut PVC coupons were sterilized simultaneously by

immersion in 1 liter of 70% ethanol (Fisher Scientific, USA) for 5 min. Ethanol was removed

from the coupons‘ surfaces by aseptically washing with 2 liters of sterile, deionized water over

the coupons in 0.5 liter aliquots. Waxed cardboard coupons (International Paper Company,

Griffin, GA) were cut to size, 24 x 90 cm, 4 mm thick using a utility knife. The coupons were

sterilized by wiping 70% ethanol over front and back side surfaces and allowed to stand 5 min.

Ethanol (70%) was removed from coupon surfaces by wiping sterile deionized water over front

and back side surfaces using a sterile Whirl Pack ―Speci-Sponge‖ sampling sponge (Nasco, Fort

Atkinson, WI). Stainless steel coupons were sterilized and washed for use throughout the study,

PVC and waxed cardboard coupons were cut for one-time use. After sterilization, coupons were

aseptically transferred to sterile metal pans for drying in a biosafety cabinet (Bio Safety

Cabinets, NuAire, Class II Type A2, Plymouth, MN).

The resuspended cocktail of Salmonella enterica was spot inoculated in 100 μl aliquots

onto dry coupon surfaces using a micropipette (Eppendorf Reference 100 µl, Precision Pipette

Inc., Atlanta, GA) and sterile tips (5-200 µl NATURAL, Dot Scientific Inc., Burton, MI). Spots

of approximately 5 μl were placed in a five-die pattern on the face of the coupons. Inoculum on

target coupons was allowed to dry for 90 min once all target coupons were inoculated.

37

Preparation of Sanitizer Spray Liquids

Sanitizers used in the study were Oxonia Active (Ecolab, Inc. St. Paul, MN), a peracetic

acid product, and Whisper V (Ecolab, Inc. St. Paul, MN), a quaternary ammonium compound

product. A control spray liquid of sterile tap water was used at standard conditions, 25°C and

20-25 ppm hardness measured by Water Hardness Test Kit (Ecolab, Inc. St. Paul, MN).

Sanitizer spray liquids intended for use in hydraulic-atomizing nozzle were prepared according

to manufacturer instructions for sanitizing food contact surfaces.

Microbiological Analysis of Samples

Sprayed target coupons were removed from the spray arc, allowed to stand in fabricated

holders for 10 min, and were aseptically placed into individual stomacher bags (VWR, USA)

containing 20 ml of DE neutralizing broth (Becton Dickinson). Target coupons were hand

massaged in stomacher bags for 30 sec. Serial dilutions were made using 9 ml 0.1% peptone

water and dilutions were spread plated on tryptic soy agar (Becton Dickinson) with 0.5% yeast

extract. Plates were incubated at 37±2°C for 24 h before enumeration using a Quebec Dark-

Field Colony Counter (Reichert Analytical Instruments, Depew, NY).

Statistical Analysis

Each experiment was replicated three times. For experiment 1, the data for mass transfer

analyzed in terms of ng tracer/coupon face deposited and μl carrier liquid/coupon face deposited.

Experiment 1 compared the three nozzle types under three different target orientations: (1)

perpendicular to incoming spray vector and front side, (2) perpendicular to spray vector and back

side, and (3) parallel to spray vector side. Analysis of variance (ANOVA) was performed under

each condition, and the Tukey multiple pair-wise test was used to compare the three nozzle

38

types. The significance level was set at p<0.05 for experiment 1. Transfer efficiency was

calculated as (measured liquid deposited/theoretical 100% liquid deposition) x 100%.

For experiment 2, the response variable was CFU/coupon log reduction Salmonella after

spray treatment. The initial population of Salmonella did not vary as consistent strains and

inoculation surface was used throughout the study. A trend line with the best possible

correlation (R2) was fitted for charged electrostatic spray application at different biocide rates

and a base line was found for hydraulic spray application at full-rate. The equivalence point was

found using the equation produced by the trend line fitted for charged electrostatic spray

application at different biocide rates.

The response variable for experiment 3 was CFU/coupon log reduction Salmonella after

spray treatment. There was a difference in the initial population of Salmonella on different

coupon surfaces, so the data were analyzed as log initial CFU/coupon – log final CFU/coupon

for each surface. Nine experimental sets were created which evaluated nozzle and sanitizer

treatments‘ effect on surface and side.

Results and Discussion

Experiment 1: Fluorometric Mass Transfer Analysis

In this experiment, three different nozzles dispensed normalized tracer mass toward

stainless steel target coupons. While tracer mass was normalized, the nozzles dispensed different

volumes of carrier liquid. The hydraulic nozzle had a larger application rate of carrier liquid at

600 ml/min to represent a flood sanitizing scenario. ESS nozzle had a lower application rate of

100 ml/min which represented a reduced-volume spray sanitizing scenario (28). In every

orientation analyzed, the air-assisted charged electrostatic nozzle had the highest tracer mass

deposited (p<0.05; Table 3.1). The air-assisted uncharged electrostatic nozzle had a significantly

39

larger amount of tracer mass deposited than the hydraulic nozzle (p<0.05; Table 3.1). The

largest difference in tracer mass deposited for charged and uncharged electrostatic nozzles can be

seen in the non-apparent orientations, back side and side.

Charged electrostatic nozzle deposited more carrier liquid than uncharged electrostatic

nozzle and hydraulic nozzle for every orientation (p<0.05; Table 3.2). The hydraulic nozzle

deposited less carrier liquid on the apparent front side surfaces than uncharged electrostatic

nozzle, but more on the non-apparent surfaces, back side and side (p<0.05; Table 3.2).

In this study the procedure of Giles and Law (14) was used with some modification for

evaluation of mass transfer analysis using fluorometry. Stainless steel targets were used. In

addition to being a commonly used food processing surface, stainless steel was selected as an

idealized surface for quantification of mass transfer, providing a material which is easily

grounded and washed for recovery of tracer. Charged electrostatic nozzle deposited a higher

mass of tracer than uncharged electrostatic nozzle and hydraulic conventional nozzle. The

deposition ratio of charged to uncharged sprays is known as the electrostatic deposition benefit

ratio. Electrostatic deposition benefits for front side targets were 9.2-fold and 1.2-fold for

hydraulic and uncharged electrostatic nozzles, respectively. Back side electrostatic deposition

benefit ratios were 29.6 and 6.1 for hydraulic and uncharged electrostatic nozzles, respectively.

Targets in an orientation parallel to the spray vector exhibited an electrostatic deposition benefit

of 6.1 and 2.9 for hydraulic and uncharged electrostatic nozzles. The largest electrostatic

deposition benefit was seen on the back side due to ―wrap-around‖ effect imparted by

electrostatic charge on droplets. Similar electrostatic deposition ratios have been documented for

various surfaces using fluorometric mass transfer analysis by Cooper and Law (9), Dai et al.

(10), Kang et al., (19) Law et al. (22, 24, 25).

40

In addition to enhanced transfer of tracer mass (tracer mass analogous to biocide mass),

electrostatically charged spray showed enhanced deposition over conventional hydraulic nozzle

of carrier liquid deposited onto stainless steel targets. Electrostatic deposition benefit of carrier

liquid was 1.6, 5.0, and 1.8 for front back and side (parallel) orientations, respectively.

Electrostatic nozzle reduced off-target spraying by reducing the total volume of liquid dispensed

from the nozzle toward the target (100 ml/min ESS vs. 600 ml/min hydraulic-atomizing) as well

as enhancing the amount of dispensed liquid actually deposited on the target surface. Carrier

liquid deposited on front side orientations was consistent with reported values from food

factories. Reported spray-mix dispense rates were 0.4–572 ml/m2, with a mean and median

dispense rate of 102 and 18.2 ml/m2 (17). Calculated spray-mix dispense rates with the setup

used in this study were 94.89 ml/m2 and 15.82 ml/m

2 for hydraulic and electrostatic nozzles,

respectively. Calculated maximum areal density of spray liquid deposited onto target coupons

for a dual pass were 8.62 µl/cm2 and 1.83 µl/cm

2, respectively, for hydraulic and electrostatic

nozzles. Charged spray deposited 1.83 µl/cm2 on front side stainless steel coupons, 100.00%

calculated transfer efficiency (Table 3.2). Uncharged sprays from the same ESS nozzle

deposited 1.53 µl/cm2 of spray liquid, significantly lower volume resulting in a transfer

efficiency of 83.61%. Hydraulic sprays deposited 1.18 µl/cm2 while having a calculated

maximum areal density of 8.62 µl/cm2, a transfer efficiency of 13.69%. Hydraulic sprays had

higher liquid volume application rates and lower areal deposition values on targets. Charged

sprays were most efficient by dispensing lower spray-mix volumes, lower dispense rates of

active ingredients, and depositing the most liquid volume on targets.

41

Experiment 2: Microbiological Equivalence Point

For peracetic acid sanitizer teatment, charged electrostatic spray of biocide produced a

logarithmic trend line. The equivalence point was 0.44 full-rate, as determined by an R2 value of

0.77 (Figure 3.1). Quaternary ammonium sanitizer treatment charged electrostatic spray of

biocide produced a linear trend line. The equivalence point was 0.70 full-rate, as determined by

an R2 value of 0.99 (Figure 3.2). At full-rate, charged sprays were more antimicrobially

effective than hydraulic sprays for both peracetic acid and quat sanitizer.

At the manufacturers‘ recommended usage rate, full-rate hydraulic spray treatment was

more effective using peracetic acid as shown by a 3.36 log CFU reduction of Salmonella

compared to a 2.26 log CFU reduction when quaternary ammonium compound was used. Full-

rate charged spray reduced Salmonella population 2.79 log CFU on front side stainless steel

coupons when quat was used as the spray liquid and 4.13 logs CFU when peracetic acid was

used. Peracetic acid was more effective than quat at every rate evaluated in the equivalence

study (p<0.001). Orth and Mrozeck found that peracetic acid was more effective than

benzalkonium chloride in 5 min at 20°C against food poisoning bacteria L. monocytogenes,

Staphylococcus aureus, and Enterobacter faecium (2). Knight and Craven found that 3% (v/v)

PAA was most effective over hypochlorite, acid-anionic, and quat sanitizers at reducing mixed

biofilm cultures from dairy factories on floor materials (21).

It has been shown in previous studies that a reduced rate of biocide applied

electrostatically is as effective as full-rate biocide applied with conventional hydraulic nozzles

(23, 40).

The stainless steel target surface was chosen as an idealized target to establish the

microbiological equivalence point because of its cleanability (4, 11, 30, 31). Peracetic acid and

42

quaternary ammonium compounds were chosen as biocides for their broad spectrum inactivation

of organisms, stability, differing liquid properties and common use in the food industry (27, 28,

34).

Experiment 3: Application of Equivalence Rate to Various Surfaces and Orientations

For stainless steel surfaces, all orientations (front, back, side) were best decontaminated

by peracetic acid santizer, followed by quat sanitizer as next best, with tap water control

removing the lowest amount of the Salmonella population (p<0.001; Table 3.3). Hydraulic

nozzle application of tap water control removed more Salmonella from the front surfaces than

tap water applied with electrostatic nozzles due to mechanical action. All sanitizer spray

treatments in this portion of the study were applied as full-rate hydraulic and equivalence rate

charged and uncharged sprays.

The front surfaces of the stainless steel targets had similar population reductions for all

spray nozzles when using the same liquid sanitizer (p<0.001; Table 3.3). Nozzle type was not

significantly different due to the apparent target orientation. However, charged spray did have

the largest population reduction of Salmonella due to the charged droplets evenly coating the

target surface (Figure 3.3; 12).

The back surfaces of the steel targets were best decontaminated by charged electrostatic

sprays, followed by hydraulic and uncharged sprays which were not different (p<0.001; Table

3.4). Electrostatic benefit was most apparent on the back surfaces of the targets. Peracetic acid

charged spray reduced populations 1.62 log CFU whereas uncharged and hydraulic sprays

reduced populations 0.07 and 0.00 log CFU, respectively (Fig. 3.4). A similar trend was seen

with quat where charged spray resulted in 1.42 log CFU reduction and uncharged, hydraulic

sprays resulted in 0.21 and 0.09 log CFU reduction each (Figure 3.4). Similar results have been

43

reported on abaxial leaf surfaces, which are analogous to back surface of targets (29). Enhanced

deposition has also been exhibited on non-apparent surfaces such as spheres and spherical

produce items (8, 20).

When the stainless steel targets were orientated parallel to the spray vector so neither the

front or back surfaces were sprayed directly (side orientation) the charged sprays were more

effective in reducing the Salmonella populations than hydraulic sprays for all liquid treatments

applied. Uncharged spray was the least effective nozzle type for decontamination of stainless

steel targets in side orientation (p<0.05; Table 3.5). Charged sprays were able to ―wrap around‖

and deposit onto unapparent surfaces in side orientation. Peracetic acid charged spray reduced

Salmonella populations on side orientation stainless steel targets 1.45 log CFU whereas

uncharged sprays reduced populations by 1.00 log CFU (Figure 3.5). Similar results were

documented by Law et al. when charged pollen sprays deposited more effectively than

uncharged sprays on targets parallel to the spray vector (26).

The front surfaces of PVC was most effectively decontaminated by peracetic acid

sanitizer, whereas tap water and quat performed similarly on front surfaces for all nozzle types

(p<0.001; Table 3.6). Charged and hydraulic nozzles performed similarly on PVC surfaces in

front orientation, and both nozzles were significantly better than uncharged electrostatic nozzle

(p<0.05; Table 3.6). Quat applied to front surface of PVC had similar population reductions for

all nozzles (Figure 3.6).

Peracetic acid on the back surfaces of PVC reduced bacterial populations better than quat,

which was more effective than tap water control (p<0.001; Table 3.7). Charged electrostatic

spray nozzle was most effective for all sanitizer applications (p<0.001), and uncharged

electrostatic spray nozzle was more effective than hydraulic nozzle (p<0.05;Table 3.7). The

44

ability to create a film on unapparent surfaces due to electrostatic droplet charging made the

charged spray more effective than uncharged and hydraulic sprays. Charged peracetic acid spray

was most effective with 1.49 log CFU reduction (Figure 3.7). Uncharged sprays were more

effective than hydraulic spray due to the pneumatic energy in the air-assisted nozzle projecting

the smaller droplets into the target area, where as the hydraulic nozzle created large droplets

which fell due to gravity once past the target face (23).

PVC in side orientation had the highest population reduction when peracetic acid

sanitizer was applied, followed by quat sanitizer. Tap water was least effective (p<0.05; Table

3.8). All three nozzles performed similarly on PVC surface in side orientation, regardless of

liquid sanitizer applied (p<0.001; Table 3.8). The most effective combination for side PVC

treatment was hydraulic spray using peracetic acid (Figure 3.8).

The population of Salmonella on the front surfaces of waxed cardboard had the greatest

level of reduction when treated with peracetic acid, while quat was equally effective as tap water

(p<0.001; Table 3.9). The hydraulic nozzle was most effective in decreasing microbial

populations on the front surfaces of waxed cardboard with charged and uncharged sprays being

similar, but less effective (p<0.001; Table 3.9). The hydraulic nozzle reduced the Salmonella

population on front surface of the waxed cardboard by 4.64 log CFU compared to 1.24 and 1.13

log CFU for charged and uncharged sprays, respectively (Figure 3.9). The waxed cardboard

surface was highly hydrophobic due to the wax film applied to the paperboard as a barrier

against water. A high volume of carrier liquid along with mechanical energy from the hydraulic

nozzle was needed to film the surface and significantly reduce the microbial population on

waxed cardboard (27, 35).

45

For the back surfaces of waxed cardboard, peracetic acid sanitizer performed

significantly better than quat sanitizer and tap water sprays which were similar (p<0.001; Table

3.10). The charged spray resulted in a greater population reduction on the back surfaces of the

waxed cardboard than uncharged and hydraulic sprays (p<0.05; Table 3.10). While the hydraulic

spray was the most effective when spraying apparent waxed cardboard surfaces, it is least

effective when spraying unapparent surfaces such as the back surface. Charged peracetic acid

spray was most effective with a 0.86 log CFU reduction compared to uncharged spray and

hydraulic spray with 0.31 and 0.09 log CFU reduction, respectively (Figure 3.10).

Peracetic acid sanitizer reduced Salmonella populations on waxed cardboard surfaces in

side orientation better than quaternary ammonium sanitizer (p<0.001) and both sanitizer

treatments were significantly better than tap water control (p<0.001; Table 3.11). Charged and

hydraulic nozzles performed similarly and resulted in higher population reductions compared to

the uncharged nozzle on waxed cardboard in side orientation (p<0.05; Table 3.11). Charged

droplets were able to be deposited on the non-apparent target due to ―wrap-around‖ effect and

hydraulic droplets were able to shear across a surface parallel to the spray vector and effectively

film over target areas.

Acknowledgements

Funding was provided by National Institute of Food and Agriculture, USDA, under

special project #2009-51110-20161 and the Georgia Agricultural Experiment Stations.

Statistical analysis was provided by Dr. Jien Chen and Dr. Jaxk Reeves (Department of

Statistics, University of Georgia).

46

References

1. Ailes, E. C., J. S. Leon, L. A. Jaykus, L. M. Johnston, H. A. Clayton, S. Blanding, D. G.

Kleinbaum, L. C. Backer, and C. L. Moe. 2008. Microbial concentrations on fresh

produce are affected by postharvest processing, importation, and season. J. Food Prot.

71:2389-2397.

2. Block, S. S. (ed.). 2001. Disinfection, sterilization, and preservation. Lippincott

Williams & Wilkins, Philadelphia, PA.

3. Behling, R. G., J. Eifert, M. C. Erickson, J. B. Gurtler, J. L. Kornacki, E. Line, R.

Radcliff, E. T. Ryser, B. Stawick, and Z. Yan. 2010. Selected pathogens of concern to

industrial food processors: infectious, toxigenic,toxico-infectious, selected emerging

pathogenic bacteria. In J.L. Kornacki (ed.), Principles of microbiological

troubleshooting in the industrial food processing environment. Springer

Science+Business Media, LLC, New York, NY.

4. BoulangePetermann, L. 1996. Processes of bioadhesion on stainless steel surfaces and

cleanability: a review with special reference to the food industry. Biofouling. 10:275-

300.

5. Brackett, R. E. 1999. Incidence, contributing factors, and control of bacterial pathogens

in produce. Postharvest Biol. Technol. 15:305-311.

6. Centers for Disease Control and Prevention. 2010. Preliminary FoodNet data on the

incidence of infection with pathogens transmitted commonly through food - 10 states,

2009. Morbidity and Mortality Weekly Report, vol. 59.

7. Centers for Disease Control and Prevention. 2010. Salmonella outbreaks. Available at:

http://www.cdc.gov/salmonella/outbreaks.html. Accessed 14 October 2010.

8. Chaim, A., M. Pessoa, and V. L. Ferracini. 2002. Pesticide efficiency deposition

obtained with electrostatic spray head for knapsack mistblower sprayer. Pesqu.

Agropecu. Bras. 37:497-501.3.

9. Cooper, S. C., and S. E. Law. 2006. Electrostatic sprays for sunless tanning of the

human body. IEEE Trans. Ind. Appl. 42:385-391.

10. Dai, Y., S. C. Cooper, and S. E. Law. 1992. Effectiveness of electrostatic, aerodynamic

and hydraulic spraying methods for depositing pesticide sprays onto inner plant regions

and leaf undersides. Trans. ASAE (Am. Soc. Agric. En.) No. 92-1094, Charlotte, NC.

11. Davies, J., C. S. Nunnerley, A. C. Brisley, J. C. Edwards, and S. D. Finlayson. 1996.

Use of dynamic contact angle profile analysis in studying the kinetics of protein removal

from steel, glass, polytetrafluoroethylene, polypropylene, ethylenepropylene rubber, and

silicone surfaces. J. Colloid Interface Sci. 182:437-443.

http://www.cdc.gov/salmonella/outbreaks.html

47

12. Evans, M. D., S. E. Law, and S. C. Cooper. 1994. Fluorescent spray deposit

measurement via light intensified machine vision. ASAE (Am. Soc. Agric. Eng.) No. 92-

6571, St. Joseph, MI.

13. Frost, A. R., and S. E. Law. 1981. Extended flow characteristics of the embedded-

electrode spray-charging nozzle. J. Agric. Engineering Res. 26:79-86.

14. Giles, D. K. and S.E. Law. 1985. Space charge deposition of pesticide sprays onto

cylindrical target arrays. Trans. ASAE (Am. Soc. Agric. Eng.) 28:658-664.

15. Helke, D. M., and A. C. L. Wong. 1994. Survival and growth characteristics of Listeria

monocytogenes and Salmonella typhimurium on stainless steel and buna-N rubber. J.

Food Prot. 57:963-968.

16. Holah, J. T., A. Lavaud, W. Peters, and K. A. Dye. 1998. Future techniques for

disinfectant efficacy testing. Int. Biodeterior. Biodegrad. 41:273-279.

17. Holah, J. T., J. H. Taylor, D. J. Dawson, and K. E. Hall. 2002. Biocide use in the food

industry and the disinfectant resistance of persistent strains of Listeria monocytogenes

and Escherichia coli. J. Appl. Microbiol. 92:111S-120S.

18. Hsu, S.-Y., C. Kim, Y.-C. Hung, and S. E. Prussia. 2004. Effect of spraying on chemical

properties and bactericidal efficacy of electrolysed oxidizing water. Int. J. Food Sci.

Technol. 39:157-165.

19. Kang, T.-G., D.-H. Lee, C.-S. Lee, S.-H. Kim, G.-I. Lee, W.-K. Choi, and S.-Y. No.

2004. Spray and depositional characteristics of electrostatic nozzles for orchard sprayers.

ASAE Annual Meeting. Paper number 041005.

20. Kim, C., and Y. C. Hung. 2007. Development of a response surface model of an

electrostatic spray system and its contributing parameters. Trans. ASABE. (Am. Soc.

Agric. Biol. Eng.) 50:583-590.

21. Knight, G. C., and H. M. Craven. 2010. A model system for evaluating surface

disinfection in dairy factory environments. Int. J. Food Microbiol. 137:161-167.

22. Law, S. E. 1978. Embedded-electrode electrostatic-induction spray-charging nozzle -

theoretical and engineering design. Trans. ASAE (Am. Soc. Agric. Eng.) 21:1096-1104.

23. Law, S. E., and S. C. Cooper. 2001. Air-assisted electrostatic sprays for postharvest

control of fruit and vegetable spoilage microorganisms. IEEE Trans. Ind. Appl.

37:1597-1602.

24. Law, S. E., S. C. Cooper, and M. A. Harrison. 2004. Electrostatic spray application of

decontaminant agents onto the human body as a bioterrorism countermeasure: process

48

development and evaluation. p. 331-336. In H. Morgan (ed.), Electrostatics 2003:

proceedings of the Electrostatics Conference of the Institute of Physics: held in

Edinburgh, UK, 23-27 March, 2003. CRC Press LLC, Boca Raton, FL.

25. Law, S. E., and M. D. Lane. 1981. Electrostatic deposition of pesticide spray onto foliar

targets of varying morphology. Trans. ASAE (Am. Soc. Agric. Eng.) 24:1441-1445, 1448.

26. Law, S. E., H. Y. Wetzstein, S. Banerjee, and D. Eisikowitch. 2000. Electrostatic

application of pollen sprays: effects of charging field intensity and aerodynamic shear

upon deposition and germinability. IEEE Trans. Ind. Appl. 36:998-1009.

27. Lelieveld, H. L. M., M. A. Mostert, J. Holah, and B. White (ed.). 2000. Hygiene in food

processing. CRC Press LLC, Boca Raton, FL.

28. Marriot, N. G., and R. B. Gravani. 2006. Principles of food sanitation. Springer

Science+Business Media, Inc., New York, New York.

29. Maski, D., and D. Durairaj. 2010. Effects of charging voltage, application speed, target

height, and orientation upon charged spray deposition on leaf abaxial and adaxial

surfaces. Crop Prot. 29:134-141.

30. Midelet, G., and B. Carpentier. 2002. Transfer of microorganisms, including Listeria

monocytogenes, from various materials to beef. Appl. Environ. Microbiol. 68:4015-

4024.

31. Moore, G., I. S. Blair, and D. A. McDowell. 2007. Recovery and transfer of Salmonella

typhimurium from four different domestic food contact surfaces. J. Food Prot. 70:2273-

2280.

32. Musgrove, M. T., D. R. Jones, J. K. Northcutt, P. A. Curtis, K. E. Anderson, D. L.

Fletcher, and N. A. Cox. 2004. Sources and risk factors for contamination, survival,

persistence, and heat resistance of Salmonella in low-moisture foods. J. Food Prot.

67:1919-1936.

33. Podolak, R., E. Enache, W. Stone, D. G. Black, and P. H. Elliot. 2004. Survey of shell

egg processing plant sanitation programs: effects on non-egg-contact surfaces. J. Food

Prot. 67:2801-2804.

34. Polat, M., H. Polat, and S. Chander. 2000. Electrostatic charge on spray droplets of

aqueous surfactant solutions. J. Aerosol Sci. 31:551-562.

35. Pordesimo, L. O., E.G. Wilkerson, A.R. Womac, and C.N. Cutter. 2002. Process

engineering variables in the spray washing of meat and produce. J. Food Prot. 65:222-

237.

49

36. Russell, S. 2003. The effect of electrolyzed oxidative water applied using electrostatic

spraying on pathogenic and indicator bacteria on the surface of eggs. Poult. Sci. 82:158-

162.

37. Sansebastiano, G., R. Zoni, and L. Bigliardi. 2007. Cleaning and disinfection procedures

in the food industry general aspects and practical applications. p. 253-280. In A.

McElhatton, and R.J. Marshall (ed.), Food safety: a practical and case study approach.

Springer Science+Business Media, LLC, New York, NY.

38. Scallan E, Hoekstra RM, Angulo FJ, Tauxe RV, Widdowson M-A, Roy SL, et al.

January 2011. Foodborne illness acquired in the United States—major pathogens.

Emerg Infect Dis [serial on the internet]. Available at:

http://www.cdc.gov/EID/content/17/1/7.htm. Accessed 31 March 2011.

39. Scherm, H., A. T. Savelle, and S. E. Law. 2007. Effect of electrostatic spray parameters

on the viability of two bacterial biocontrol agents and their deposition on blueberry

flower stigmas. Biocontrol Sci. and Technol. 17:285-293.

40. Starrs, C. 28 February 2009. New applications help company with patented spray

system grow. Available at:

http://www.onlineathens.com/stories/030109/bus_399637493.shtml. Accessed 14

October 2010.

41. United States Department of Agriculture. 2010. Foodborne illness cost calculator:

Salmonella. Available at:

http://www.ers.usda.gov/Data/FoodborneIllness/salm_Intro.asp. Accessed 14 October

2010.

42. Washington Metropolitan Area Transit Authority. 2009. ESS - D.C. Metro prepares for

Swine Flu. Available at: http://www.youtube.com/watch?v=l3kmRQx8m3A. Accessed

14 October 2010.


http://www.onlineathens.com/stories/030109/bus_399637493.shtml

http://www.ers.usda.gov/Data/FoodborneIllness/salm_Intro.asp.%20Accessed%2014%20October%202010

http://www.youtube.com/watch?v=l3kmRQx8m3A

50

Chapter 4

CONCLUSION

This study showed air-assisted induction-charged sprays of sanitizers are an effective

means to decontaminate food contact surfaces. Compared with conventional hydraulic-nozzle

sanitizer application, charged electrostatic sprays reduced both the amount of biocide used and

carrier liquid applied to the target area while achieving the same or greater population reduction

of Salmonella. Peracetic acid sanitizer was more effective than quaternary ammonium

compound sanitizer in all spray applications. Electrostatically charged sprays are an effective

sanitizer application method which can be used in conjunction with adequate sanitation programs

in the food processing environment.

51

Table 3.1. Tracer deposition on stainless steel coupons normalized to equal tracer mass

dispensed toward target coupons using three spray treatments (Hydraulic: hydraulic

nozzle dispensing 600 ml of sanitizer spray/min, Uncharged: uncharged electrostatic


nozzle dispensing 100 ml of sanitizer spray/min). Front side, back side: n = 12.

Side: n = 24.

Coupon target

Tracer deposited (ng/cm2)

Hydraulic Uncharged Charged

Front side 592.10a

4590.20b

5490.50c

Back side 55.85a

271.39b

1655.10c

Side 340.14a

730.76b

2091.30c

abc Means within a row with different superscripts differ (p<0.05).

52

Table 3.2. Carrier liquid deposition on stainless steel coupons using three spray treatments

(Hydraulic: hydraulic nozzle dispensing 600 ml of sanitizer/min, Uncharged:

uncharged electrostatic nozzle dispensing 100 ml of sanitizer/min, Charged: charged

electrostatic nozzle dispensing 100 ml of sanitizer/min). Front side, back side: n = 12.

Side: n = 24.

Coupon target Liquid deposited, μl/cm

2 (calc. transfer efficiency

1)

Hydraulic Uncharged Charged

Front side 1.18a (13.69%) 1.53

b (83.61%) 1.83

c (100.00%)

Back side 0.11b

0.09a

0.55c

Side 0.42b

0.30a

0.75c

1 (Experimental deposition density/theoretical deposition density) x 100%


53

Figure 3.1. Effect of relative rate of peracetic acid sanitizer on population density of

Salmonella on stainless steel coupons (20.7 cm2) applied as a low volume



y = 1.347ln(x) + 4.458, (R2 = 0.77). The dashed line and open symbol with error

bars represent the mean log CFU reduction per coupon face when peracetic acid is

applied as a hydraulic spray at full rate. n = 12.

0.00

1.00

2.00

3.00

4.00

5.00

0.00 0.25 0.50 0.75 1.00

logC

FU r

ed

uct

ion

pe

r co

up

on

fac

e

Relative rate of peracetic acid

54

Figure 3.2. Effect of relative rate of quaternary ammonium sanitizer on population density of

Salmonella on stainless steel coupons (20.7 cm2) applied as a low-volume



y = 1.934x + 0.901, (R2 = 0.99). The dashed line and open symbol with error bars

represent the mean log CFU reduction per coupon face when peracetic acid is

applied as a hydraulic spray at full rate. n = 12.

0.00

1.00

2.00

3.00

4.00

0.00 0.25 0.50 0.75 1.00

logC

FU r

ed

uct

ion

pe

r co

up

on

fac

e

Relative rate of quaternary ammonium compound

55

Table 3.3. Reduction of Salmonella populations on the front surfaces of stainless steel coupons

after treatment with peracetic acid, quaternary ammonium compounds, and tap water

using three spray treatments. Biocide dispensed full-rate hydraulic vs. equivalence-

rate charge and uncharged. n = 12.

Spray Liquid

Log reduction (CFU/coupon)1

Peracetic acid 3.62z

Quaternary ammonium compounds 1.98y

Tap water 0.24x

1 Values expressed as least square means of log reduction CFU/coupon

xyz Means within a column with different superscripts differ (p<0.001).

56

Figure 3.3. Population reduction of Salmonella on the front surface of stainless steel coupons







Detection limit: 1.9 log CFU/coupon face. n = 12.

0.04

2.21

3.82

0.02

1.79

3.41

0.68

1.97

3.61

-1.0

0.0

1.0

2.0

3.0

4.0

5.0

Log

CFU

re

du

ctio

n p

er

cou

po

n f

ace

Charged

Uncharged

Hydraulic

Peracetic acid Tap waterQuat

Spray liquid

57

Table 3.4. Reduction of Salmonella populations on the back surfaces of stainless steel coupons


using three spray treatments (Hydraulic: hydraulic nozzle dispensing 600 ml of

sanitizer spray/min, Uncharged: uncharged electrostatic nozzle dispensing 100 ml of

sanitizer spray/min, Charged: charged electrostatic nozzle dispensing 100 ml of

sanitizer spray/min). Biocide dispensed full-rate hydraulic vs. equivalence-rate charge

and uncharged. n = 12.

Spray Liquid

Log Reduction1

Peracetic acid 1.09b

Quaternary ammonium compounds 0.52c

Tap water -0.17a

Spray Nozzle Log Reduction

Hydraulic 0.07x

Uncharged 0.17x

Charged 1.20y

1 Values expressed as least square means of log reduction CFU/coupon.


xy Means within a row with different superscripts differ (p<0.001).

58

Figure 3.4. Population reduction of Salmonella on the back surface of stainless steel coupons


using three spray treatments (Hydraulic: hydraulic nozzle dispensing 600 ml of

sanitizer spray/min, Uncharged: uncharged electrostatic nozzle dispensing 100 ml of

sanitizer spray/min, Charged: charged electrostatic nozzle dispensing 100 ml of

sanitizer spray/min). Biocide dispensed full-rate hydraulic vs. equivalence-rate

charge and uncharged. Initial population: 106 CFU/coupon face. Detection limit: 1.9

log CFU/coupon face. n = 12.

1.62

1.42

0.37

0.07

0.21

0.060.000.09 0.09

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

1.8

2.0

Log

CFU

re

du

ctio

n p

er

cou

po

n f

ace

Charged

Uncharged

Hydraulic


Spray liquid

59

Table 3.5. Reduction of Salmonella populations on the side surfaces of stainless steel






hydraulic vs. equivalence-rate charge and uncharged. n = 12.

Spray Liquid

Log Reduction1

Peracetic acid 1.31c

Quaternary ammonium compounds 0.82b

Tap water -0.91a


Hydraulic 0.44y

Uncharged 0.18x

Charged 0.60z



xyz Means within a row with different superscripts differ (p<0.001).

60

Figure 3.5. Population reduction of Salmonella on the side surfaces of stainless steel coupons


water using three spray treatments (Hydraulic: hydraulic nozzle dispensing 600 ml

of sanitizer spray/min, Uncharged: uncharged electrostatic nozzle dispensing 100

ml of sanitizer spray/min, Charged: charged electrostatic nozzle dispensing 100 ml

of sanitizer spray/min). Biocide dispensed full-rate hydraulic vs. equivalence-rate

charge and uncharged. Initial population: 106 CFU/coupon face. Detection limit: 1.9

log CFU/coupon face. n = 24.

1.45

1.28

0.27

1.00

0.42 0.31

1.47

0.77

0.26

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

1.8

Log

CFU

re

du

ctio

n p

er

cou

po

n f

ace

Charged

Uncharged

Hydraulic


Spray liquid

61

Table 3.6. Reduction of Salmonella populations on the front surfaces of polyvinyl chloride

coupons after treatment with peracetic acid, quaternary ammonium compounds, and

tap water using three spray treatments (Hydraulic: hydraulic nozzle dispensing 600

ml of sanitizer spray/min, Uncharged: uncharged electrostatic nozzle dispensing 100

ml of sanitizer spray/min, Charged: charged electrostatic nozzle dispensing 100 ml of


and uncharged. n =12.

Spray Liquid

Log Reduction1

Peracetic acid 2.86a


Tap water 0.79b


Hydraulic 1.57y

Uncharged 1.15x

Charged 1.70y


ab Means within a row with different superscripts differ (p<0.001).


62

Figure 3.6. Population reduction of Salmonella on the front surfaces of polyvinyl chloride




100 ml of sanitizer spray/min, Charged: charged electrostatic nozzle dispensing 100

ml of sanitizer spray/min). Biocide dispensed full-rate hydraulic vs. equivalence-

rate charge and uncharged. Initial population: 106 CFU/coupon face. Detection

limit: 1.9 log CFU/coupon face. n = 12.

3.60

0.800.70

2.21

0.74

0.49

2.75

0.79

1.17

0.0

1.0

2.0

3.0

4.0

Log

CFU

re

du

ctio

n p

er

cou

po

n f

ace

Charged

Uncharged

Hydraulic


Spray liquid

63

Table 3.7. Reduction of Salmonella populations on the back surfaces of polyvinyl chloride


and tap water using three spray treatments1 (Hydraulic: hydraulic nozzle




hydraulic vs. equivalence-rate charge and uncharged. n = 12.

Spray Liquid

Log Reduction1



Tap water -0.09a


Hydraulic -0.08x

Uncharged 0.16y

Charged 0.68z



xyz Means within a row with different superscripts differ (p<0.001).

64

Figure 3.7. Population reduction of Salmonella on the back surfaces of polyvinyl chloride








1.49

0.48

0.07

0.220.17

0.09

0.000.05

0.16

-0.2

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

1.8

Log

CFU

re

du

ctio

n p

er

cou

po

n f

ace

Charged

Uncharged

Hydraulic


Spray liquid

65

Table 3.8. Reduction of Salmonella populations on the side surfaces of polyvinyl chloride







Spray Liquid Log reduction (CFU/coupon)1

Peracetic acid 0.51z

Quaternary ammonium compounds 0.26y

Tap water 0.16x

1 Values expressed as least square means of log reduction CFU/coupon

xyz Means within a column with different superscripts differ (p<0.001).

66

Figure 3.8. Population reduction of Salmonella on the side surfaces of polyvinyl chloride








0.46

0.28 0.240.31

0.180.12

0.77

0.32

0.13

-0.2

0.0

0.2

0.4

0.6

0.8

1.0

Log

CFU

re

du

ctio

n p

er

cou

po

n f

ace Charged

Uncharged

Hydraulic


Spray liquid

67

Table 3.9. Reduction of Salmonella populations on the front surfaces of waxed cardboard







Spray Liquid

Log Reduction1


Quaternary ammonium compounds 0.21a

Tap water 0.22a


Hydraulic 1.67y

Uncharged 0.52x

Charged 0.58x




68

Figure 3.9. Population reduction of Salmonella on the front surfaces of waxed cardboard








1.24

0.24 0.27

1.13

0.260.17

4.64

0.140.24

0.0

1.0

2.0

3.0

4.0

5.0

Log

CFU

re

du

ctio

n p

er

cou

po

n f

ace

Charged

Uncharged

Hydraulic


Spray liquid

69

Table 3.10. Reduction of Salmonella populations on the back surfaces of waxed cardboard







Spray Liquid

Log Reduction1


Quaternary ammonium compounds 0.19a

Tap water 0.18a


Hydraulic 0.13x

Uncharged 0.19x

Charged 0.47y




70

Figure 3.10. Population reduction of Salmonella on the back surfaces of waxed cardboard








0.86

0.21

0.340.31

0.19

0.07

0.090.17

0.13

-0.2

0.0

0.2

0.4

0.6

0.8

1.0

1.2Lo

g C

FU r

ed

uct

ion

pe

r co

up

on

fac

e

Charged

Uncharged

Hydraulic


Spray liquid

71

Table 3.11. Reduction of Salmonella populations on the side surfaces of waxed cardboard







Spray Liquid

Log Reduction1



Tap water 0.24a


Hydraulic 0.53y

Uncharged 0.37x

Charged 0.56y




72

Figure 3.11. Population reduction of Salmonella on the side surfaces of waxed cardboard








0.88

0.52

0.27

0.54

0.37

0.20

0.83

0.49

0.25

0.0

0.2

0.4

0.6

0.8

1.0

1.2

Log

CFU

re

du

ctio

n p

er

cou

po

n f

ace

Charged

Uncharged

Hydraulic


Spray liquid

73

Chapter 5

APPENDIX A

74

Figure A.1. Food Science and Technology spray apparatus and target chamber – side view.

75

Figure A.2. Food Science and Technology spray target chamber – frontal view.

76

Figure A.3. Spray arm pathway dimensions.

Arm radius = 100 cm, arc of sweep of nozzle = 120°

77

Calculation of application rate of liquid dispensed from spray nozzle

Number of passes = 2

Time for 2 passes (T) = 6 s

ESS nozzle liquid flow rate (VL) = 100 ml/min

Hydraulic nozzle liquid flow rate (VL) = 600 ml/min

Area of spray path (Ap) = (Πr2 outside circle – Πr

2 inside circle)(1/3circle)

= (Π45in2– Π33in

2)(1/3circle)

= 2940.53 in2(1/3circle)

= 980.177 in2 = 0.6323 m

2

Volume of liquid dispensed in spray path (Vp) = (VLT)/Ap

Vp hydraulic = 60 ml/0.6323 m2

= 94.89 ml/m2

Vp ESS = 10 ml/0.6323 m2

= 15.82 ml/m2

78

Calculation of estimated spray interception by target coupon

ESS nozzle spray pattern diameter at target when 30 in. from nozzle face (D) = 12 in = 1 ft.

Area of ESS nozzle circular spray pattern (AS) = ΠD2/4 = Π1

2 ft /4 = 0.7854 ft

2

ESS VL/ AS = (100 ml/min)/(0.7854 ft2) = 0.002283 (ml/s)/cm

2

Hydraulic nozzle spray pattern dimensions at target when 16.5 in. from nozzle face (LxW) = 12

in. x 4 in.

Area of hydraulic nozzle spray pattern (AS) = LxW = 1 ft x 0.33 ft = 0.33 ft2

Hydraulic VL/ AS (600ml/min)/(0.3333 ft2) = 0.03229 (ml/s)/cm

2

Target coupon area (At) = 24 mm x 90 mm = 21.6 cm2

Θ = (120°)(2Π radians/360°) = 2.094 radians/sweep

w = average angular velocity of nozzle = (dΘ/dt) = (arc length traveled)/(T observed time)

Vt = tangential velocity of nozzle past targets = (radius arm x w)

Digital timer on robotic arm clocks the total time in milliseconds for dual pass = t seconds

Average w = (2 x 2.094 radians) / T seconds

Average vt = (3.25 ft)((2 x 2.094 radians) / T seconds) = 13.611 ft/s

Assume that due to gravitational effects and time lost at ends of arc for stop/reverse de-

accelerating/accelerating, the effective tangential velocity for nozzle actually traveling in target-

holding arc sector is 10% greater than average Vt.

Effective Vt = (1.10)(average vt) = (1.10)(13.611/t) ft/s = 1.4972/T ft/s

Time for target coupon to pass through spray cloud, seconds each pass (Tt) = (D/Vt)

ESS Tt = (1 ft/(14.972 ft/6 s)) = 0.4007 s

Hydraulic Tt = (0.33 ft/(14.972 ft/6 s)) = 0.1334 s

Spray volume intercepted by target coupon per pass = (VL/ AS)(Tt)(At), ml

Time (T) for dual pass = 6000 milliseconds = 6 seconds

79

Table A.1. Calculated spray interception by target coupon.

Calculations for robotic arm

dual pass, T = 6 seconds

ESS nozzle

(100 ml/min)

Hydraulic nozzle

(600 ml/min)

Vt , velocity to traverse a

target (m/s) 2.495 2.495

Tt, time for target coupon to

pass through spray (s) 0.4007 0.1334

(VL/ AS)Tt, intercepted spray

flux density per pass (ml/ft2)

0.8505 4.0032

(VL/ AS)Tt, intercepted spray

flux density per pass (ml/cm2)

0.000915 0.00431

VL,t, volume of spray liquid

intercepted by coupon per pass

(µl)

19.760 93.042

2 x VL,t, volume of spray

liquid intercepted by coupon

per dual pass (µl)

39.520 186.083

Maximum areal density of

liquid deposited for dual pass

(µl/cm2)

1.8296 8.6150

Maximum areal density of

liquid deposited for dual pass

(ml/m2)

18.296 86.150

80

Design specifications for electrostatic spraying system

A spray liquid flow rate of 100 mL/min liquid dispensed was held constant using a 0-120

ml/min flow meter (Key Instruments, Trevose, PA). A valve between reservoir and flow meter

facilitated simple on/off control of liquid. Air pressure supplied to the nozzle was 207 kPa (30

psig). Nozzle-to-target spacing was 0.76 m (30 in.) with a nozzle orientation of 0° off horizontal

for all experiments. The spray cloud issuing from the electrostatic nozzle diverged to a width of

0.30 m (12 in.) at the face of the target.

Charge-to-mass ratio calculation:

-12 µA measured on spray cloud = -12 µC/s (1A = 1 C/s)

100 ml/min ESS nozzle flow rate = 100 g/60s (100 ml water = 100 g, 1 min = 60 s)

-12 µC/s x 60 s/100 g = -7.2 µC/g

6 µC/g x (1000/1000) = -7.2 mC/kg

Design specifications for conventional spray system

Spraying Systems Co. Teejet Even Flat Spray Tip TP40015E was used with a slotted

strainer part 451-NY-20 as the hydraulic, conventional spray system. A liquid flowrate of 600

ml/min was achieved at 296 kPa (43 psig) hydraulic pressure. Nozzle-to-target spacing was 0.42

m (16.5 in.) yielding a spray pattern swath width of 0.30 m (12 in.) at the face of the target. The

flat spray tip was positioned in a vertical spray pattern.

The conventional spray volume to electrostatic spray volume ratio was:

Nozzle spray ratio = 600 ml/min (conventional volume) / 100 ml/min (electrostatic volume) =

6.0

81

Electrostatic spray system set-up procedure

1) Attach electrostatic nozzle to spray arm, connecting the liquid inlet, air inlet, grounding

wire, and power source.

2) Connect air supply to compressed air line.

3) Turn on Alkar chamber exhaust fan and main blower.

4) Set atomizing air pressure to 30 psig and allow air to flow through nozzle for 5 min.

5) Fill spray reservoir with sterile tap water, turn on magnetic stir bar below reservoir.

6) Open valve to reservoir to start spraying liquid, adjust flow rate on flowmeter.

7) Turn on electrical equipment.

a. If treatment is charged, turn on Lambda power supply, Venus voltage amplifier,

and digital multimeters.

b. If treatment is uncharged, do not turn on Lambda power supply or Venus power

amplifier.

8) Set Lambda power supply to 9.65 Vdc and ensure the output on the multimeter reads

1200 Vdc output from the Venus.

9) Ensure that an ionization current of -10 μA is observed for the charged nozzle or 0 μA is

observed for uncharged nozzle using ionization probe placed 2-3 cm from nozzle orifice

on the center line.

10) Remove ionization probe from holder and place a safe distance from the spray path.

11) Empty remaining tap water from reservoir.

12) Fill reservoir with test solution and place targets in arc holder for treatments.

13) After treatments, triple rinse reservoir with sterile tap water. Run 400 ml sterile tap water

through tubing and nozzle to clean.

82

14) The nozzle is cleaned by unscrewing its outer cap and carefully and wiping with a

Kimwipe, including the inside bore.

Conventional hydraulic spray system set-up procedure

1) Connect pressurized water supply to Dayton Twin Piston water pump.

2) Turn on Alkar chamber exhaust fan and main blower.

3) Fill reservoir with sterile tap water, turn on magnetic stir bar below.

4) Turn on the pump‘s Dayton Industrial Motor with hydraulic line open to air.

5) Allow sterile tap water to flow through nozzle-less spray arm until debris clears.

6) Attach hydraulic nozzle to spray arm, ensuring the flat tip is oriented vertically with

respect to the spray arc.

7) Turn on Dayton Industrial Motor, set hydraulic pressure at 43 psig beginning at 0 psig

and working upward.

8) Empty remaining tap water from reservoir.

9) Fill reservoir with test solution and place targets in arc holder for treatments.

10) After treatments, triple rinse reservoir with sterile tap water. Run 4 liters of sterile tap

water through tubing to clean. Ensure that all water is cleaned from pump to prevent rust.

11) The nozzle and strainer are cleaned by unscrewing and wiping with a Kimwipe.

83

Figure A.4 Input-output operational characteristics of Venus dc-to-dc high-voltage converter.

Input voltage source: Lambda Model LLS6108 vs. output volts measured by:

Extech Digital Multimeter Model EX410

y = 128.3134x - 40.1286

R2 = 0.99

0

200

400

600

800

1000

1200

1400

0.00 1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00

DC Input Volts

DC

Ou

tpu

t V

olt

s

84

Specifications of fluorescent tracer material

Day-Glo Corporation (Cleveland, OH) GT-series based on a thermoset resin with the following

properties:

Color: Blaze orange GT-15N

Specific gravity: 1.37

Average particle size: 4-5 µm

Decomposition point: 195°C

Oil absorption: 51 lb. oil/100 lbs.

Bulking value: 0.0875 gal/lb.

General solubility: Insoluble in water, hydrocarbons, and many common solvents

Lot #: 5549B

Calibration of Turner Model 450 Digital Fluorometer

The procedure from Durham Ken Giles 1983 M.S. thesis was used to calibrate the fluorometer

(28). For the calibration and throughout the testing, the fluorometer was operated on the 200 gain

setting, with a fixed span.

The regression line produced from the calibration was found to be: y = 0.6978x + 43.2

Where:

x = tracer concentration of liquid sample (μg/liter solution)

y = net fluorometer reading (fluorometer units)

Equation R2 value = 0.9984

Background reading on wash solution was found to be 43.2 fluorometer units. The polycarbonate

wash vessel did not affect the background reading.

85

Figure A.5. Fluorescent tracer deposition on stainless steel coupons as observed under

ultraviolet lamp. A: ESS ON front, B: ESS ON back, C: ESS OFF front, D: ESS

OFF back, E: Hydraulic front, F: Hydraulic back

86

Manufacturers’ instructions for biocide dilution in water

Whisper V for sanitizing equipment in food processing plants and dairies: 1.0-2.67 oz per 4

gallons (29.6-79.0 ml per 15.1 liters) of 400 ppm hard water.

Oxonia Active for sanitizing food contact surfaces: 1.0-1.4 oz per 4 gallons (29.6-41.4 ml per

15.1 liters) of water (0.20-0.28% v/v concentration).

Preparation of sanitizers spray-mix for use in microbiological reduction equivalence study

1) Clean six 1 liter, one 2 liter, and two 4 liter Erlenmeyer flasks using Micro90 cleaner

followed by a triple rinse of tap water and a triple rinse of DI water.

2) Create a 2 liter stock solution

a. 1,200 ppm quat: Add 24 ml of Whisper V quaternary ammonium compound sanitizer to

1,976 ml of sterile tap water in 2 liter flask.

b. 1.61% (v/v) biocide: Add 32.76 ml of Oxonia Active acid liquid santizer to 2,000 ml of

sterile tap water in 2 liter flask.

3) Combine stock solution with the following volumes of sterile tap water in 1 or 4 liter

flask to create equivalence point test concentrations.

4) Stir each sanitizer suspension using a sterile magnetic stir bar on magnetic stir plates for

at least 5 min prior to use.

87

Table A.2. Sanitizer mixing volumes for equivalence point test concentrations.

Sanitizer treatment Stock (ml) Water (ml) Total volume (ml)

Hydraulic control 0 3000 3000

Hydraulic full rate 500 2500 3000

ESS control 0 500 500

ESS full rate 500 0 500

ESS ½ 250 250 500

ESS ¼ 125 375 500

ESS 1/6 83.3 416.7 500

ESS 1/8 62.5 437.5 500

88

Calculation of equivalence point for peracetic acid sanitizer

Fitted trend line of charged front side application to stainless steel targets:

y = 1.3472ln(x) + 4.458

Where y = population reduction in log CFU/coupon

Where x = relative rate of peracetic acid

Full-rate hydraulic front side application to stainless steel targets:

y = 3.36 log CFU/coupon

Equivalence rate:

3.36 = 1.3462ln(x) + 4.458

x = 0.44

Calculation of equivalence point for quaternary ammonium sanitizer

Fitted trend line of charged front side application to stainless steel targets:

y = 1.934x + 0.901

Where y = population reduction in log CFU/coupon

Where x = relative rate of quaternary ammonium

Full-rate hydraulic front side application to stainless steel targets:

y = 2.26 log CFU/coupon

Equivalence rate:

2.26 = 1.934x + 0.901

x = 0.70

89

Preparation of sanitizers for use in microbiological assessment of varying surfaces and

target orientations

1) Clean one 1 liter, one 2 liter, and one 4 liter Erlenmeyer flasks using Micro90 cleaner

followed by a triple rinse of tap water and a triple rinse of DI water.

2) Create a 2 liter stock solution

a. 1,200 ppm quat: Add 24 ml of Whisper V quaternary ammonium compound sanitizer

to 1976 ml of sterile tap water in 2 liter flask

b. 1.61% (v/v) biocide: Add 32.76 ml of Oxonia Active acid liquid sanitizer to 2,000 ml

of sterile tap water in 2 liter flask

3) Combine stock solution with sterile tap water in 1 or 4 liter flask at equivalence rates.

4) Stir each sanitizer suspension using a sterile magnetic stir bar on magnetic stir plates for at

least 5 min prior to use.

90

Table A.3. Sanitizer mixing volumes for microbiological assessment of varying surfaces and

target orientations.

Sanitizer treatment Stock (ml) Water (ml) Total volume (ml)

Hydraulic control 0 3000 3000

Hydraulic full rate 500 2500 3000

ESS control 0 500 500

ESS equivalence rate, quat 350 150 500

ESS equivalence rate, acid 220 280 500

91

Figure A.6. Spot inoculation of target coupons in BSL2 biosafety cabinet.

92

Figure A.7. Spot inoculated waxed cardboard target coupons.

93

Figure A.8. Spot inoculated stainless steel target coupons.

94

Figure A.9. Spot inoculated PVC target coupons.

95

Figure A.10. Target coupon orientation with respect to incoming spray vector.

Date post:	03-Mar-2021
Category:	Documents
Upload:	others
View:	1 times
Download:	0 times

VALIDATION OF ELECTROSTATIC SPRAY AS A LOW-VOLUME ... · Piansay, Sherre Chambliss, and Yanjie...

Documents