™™™™
Raaappiddd MMMiiccrrooobbiaal Routine Monitoring in Fuueeelsss aannndd Fuuel Systems
> INSTRUCTION NOTE #1
OCTOBER 2009
Aqua-tools26, rue Charles Edouard Jeanneret78300 Poissy - France Tel : + 33 (0)1 39 75 02 20 Fax : + 33 (0)1 39 75 08 28 [email protected] www.aqua-tools.com
QQQGGGGOOOOO---MMMM™™™™™::: rrraaappppiiiddd mmmicccrrooobbiaaall mmoonniitooorringg inn fuuuueeelllss aannndddd ffuuuueeell sssyyyssteeemmmms
Fuel biocontamination has existed for many years. This contamination causes fi lter clogging, microbial induced corrosion (MIC), system fouling and changes in fuel properties (accelerated instability and increased corrosivity). These problems are due to a wide diversity of microorganisms: bacteria, protozoa, yeasts and moulds which are equally hazardous. Because of this, all of them should be taken in account and properly quantifi ed in order to effi ciently fi ght fuel contamination. This phenomenon is also amplifi ed since biofuels are developed and added in common fuels, because biofuels are a better environment for microorganisms than fossil fuels.Control of biocontamination needs to be performed in fuel, water and biofi lm. The fuel-water interface is nutrient rich. Microbes growing at this interface can draw nutrients from the fuel, while remaining in an aqueous environment. Not surprisingly, the population density of microbes at the interface can be 100,000 times greater than that in the overlying fuel and 10,000 times that in the underlying water. The aqueous phase generally contains 10 times more microbes than the fuel phase. When contamination is high, biofi lm is developing on the tank surface, and gives a good environment to the microbes. Release of biofi lm increases fuel biocontamination and spreads the biofi lm all over the system.QGO-M™ is the perfect tool to control all kinds of fuel contamination and provides an early warning of microbial growth. Used as a routine test, it allows to determine “Warning” and “Action” levels and to act before contamination becomes harmful for terminal tanks, retail sites tanks and engines.
QQQGGGGOOOOO---MMMMM™™™™™ vvssss CCCCooonnnnvvveennttionnaaall CCuulltuurrreeConventional microbiological analyses measure only a fraction of total microbes and require few days for feedback. Contrary to the conventional culture tests, QGO-M™ reveals all organisms including dormant microbes, in a few minutes.
When culture tests only provide a vague and late overview of the contamination, ATP tests allow a quick action before the apparition of danger for the system. Fuels samples (and fuel systems in general) are indeed often ill-suited for microbiological development: the biomass is inhibited and not prone to develop and breed colonies on a growth media which is, by defi nition, designed to favor one specifi c kind of microorganism.
TTThhheeee AAAAAuuddddiit PPPPrrroocccceeeessssss aaatttt DDDDeeepppppoooottssss,, RRReeeettaaaaiill sssiteeessQGO-M™ can be used for routine survey tests, performed in fuel tanks, gas stations and engines. The very easy sampling protocol allows the operator to follow the biomass evolution quickly after treating fuel with chemical treatment, as conventional biocide and/or with non chemical treatment as FuelMag© technology (within hours and/or days). Obviously, all biocides are compatible with QGO-M™.
Quench-Gone Organic Modifi ed™Rapid Microbial Routine Monitoring in Fuels and Fuel Systems
QQQGGGGGOOOOO--MMMM™™™™™ ttteeessssttt kkkitQuench-Gone Organic Modifi ed test kits are based on the measurement of ATP (Adenosine Triphosphate). ATP is the primary energy carrier for all life. Therefore, QGO-M™ measures all living cells.
ATP in presence of luciferase produces light. Since a luminometer measures light and when there is more light measured, there is more ATP.From start to fi nish, most sample analyses take 3 minutes allowing real-time preventive and corrective action.QGO-M™ kit quantifies only ATP issued from living microorganisms (intracellular ATP, called cATP). All results are calibrated and delivered in pg/ml or Equivalent Microorganisms/ml.
WWWWWWhhhheeeerrrreee ttoooo uuussseee tthhhe kkiit??QGO-M™ test kit has a wide range of action. Since it is easy to use and gives very quick results, it can be used to:• Estimate viable microbial biomass• Monitor fuel system• Audit and microbial investigation• Validate chemical and non-chemical treatment effi ciency
Use QGO-M™ to investigate and quantify total microbiological activity in: • Biofuels, Fuels for automotive, marine, Diesel Locomotives• Upstream, downstream and midstream petroleum activities• Hydrocarbon diluents, Lubricants, Crude oil• Water associated• Polymer emulsions
QGO-M™ kit content for 100 tests
Defi nition
Luminase Dropper Luciferase Enzyme Reagent, 5mL
UltraCheck 1 Dropper 1 ng ATP/mL Standard, 5mL
UltraLyse 7 Bottle cATP Extraction Reagent, 125mL
UltraLute Bottle cATP Dilution Reagent, 500mL
LumiClean Bottle Filter Wash Reagent, 500mL
• Blocked fi ltersShutdown dispensersCustomer disruption
• Tank maintenanceSignifi cant spendReduced service levelCustomer disruption
• Unhappy customersat fi lling station
Quench-Gone Organic Modifi ed™Rapid Microbial Routine Monitoring in Fuels and Fuel Systems
QGO-M™ kit: Feature/Function 2nd Gen
Compatible with BioFuels e.g. Ethanol, Biodiesel? Yes
Complete ATP extraction/recovery? Yes (>90%)
Convert result to [ATP]/mL? Yes
Same protocol for fuel phase and water phase test? Yes
Time required to complete analysis for Cellular ATP? ~5 min
Number of tests required to measure Cellular ATP? 1
Volume required for test? ≤ 20mL
Sample can be used for testing other parameters? Yes
Sample can be returned to the system after testing? Yes
> Use QGO-M™ and reduce frequency of
Quench-Gone Organic Modifi ed™Rapid Microbial Routine Monitoring in Fuels and Fuel Systems
TTTTeeeesssttt kkkkittt iiinnnsssssttrrruuuccctiooonnsss
SAMPLING RECOMMENDATIONS
QGO-M™ provides a very sensitive and precise measurement of the biomass activity. As microorganisms are present everywhere, extreme care must be taken by the operator to avoid external biological contamination.
• Clean jars, without external water or any other pollutantmust be used. For the same reason, samples must bekept in good conditions in a hermetic container.
• Ideally, samples must be tested quickly after collection(within 4 hours) as microorganisms are highly perishable. If it is not possible, store the samples in insulated containers at 5° to 10°C.
• Rinse container with alcohol by fi lling it to ¼, close it and shake it during 30 seconds.
• To avoid cross-contamination, all surfaces in contactwith the sample must be disinfected with alcohol 70°(methanol, ethanol or isopropanol)
• After collecting sample, clean the container and wipe out all dust and debris…
• QGO-M can be used even any fuel additive is present.• Do not touch the surface of pipette tips.
• Typical volume used for Fuel = 20mL• Typical volume used for Bottom Water = 5mL
NOTES ON SAMPLING PROCEDURES
Microbes are not distributed uniformly throughout fuel system. Consequently, the probability of detecting microbial contamination depends substantially on the type of sample and the location within the fuel system from which the sample is collected.Collect tap or thief sample to obtain product condition (bottom third for the thief sample if possible).To determine the product biodeterioration potential, try to collect bottom water or fuel-water interface: microbes are more present in these environments.Try to take surface samples to determine system component biodeterioration (corrosion coupons, valve components, fi lter media…). In order to have enough fuel to test, sample at least 30 ml.
More informations on sampling procedures, point and storage sampling described: ASTM D 7464 – 08 norm: “Standard Practice for Manual Sampling of Liquid Fuels, Associated Materials and Fuel System Components for Microbiological Testing.”
Partners
Quench-Gone Organic Modifi ed™Rapid Microbial Routine Monitoring in Fuels and Fuel Systems
QGGOO--MMM™™™ iinntteerrpprreettaatttiooonn gguiiiddelliinnesss foor Fueel & WWaterr
ACCEPTABLE
Fuel: < 10 pg ATP/ml
Water: < 100 pg ATP/ml
• Routine monitoring must be continued
• After many “low contamination” results, test frequency can be reduced
• Drain the tank in case of water presence
• If tested water appears contaminated, increase monitoring frequency of fuel to ensure fuel remains unaffected.
PREVENTIVE ACTION REQUIREDFuel: 10-100 pg ATP/ml
Water: 100-1000 pg ATP/ml
• Drain water tank more often and try to remove fuel-water interface
• Improve settling protocols• Test again within a month• Increase frequency of routine
monitoring if successive results appear contaminated
• If water was not contaminated, suspect import of contaminated fuel
• If tested water appears contaminated, increase monitoring frequency of fuel to ensure fuel remains unaffected.
HEAVY CONTAMINATIONFuel: > 100 pg ATP/ml
Water: > 1000 pg ATP/ml
• Retest immediately • Investigate fuel quality by tests of bulk
fuel samples (e.g. suction level, loading rack) and check quality of receipts.
• Consider treatment if fuel quality might be affected
• In case of sludge presence, clean the tank before biocide treatment.
• Drain water tank more often and try to remove fuel-water interface
• Improve settling protocols• Retest after treatment• Increase frequency of routine monitoring
✔ !
QGO-M™ test kits can be used to audit microbial quantity to reveal differences at different process locations in an effort to quickly assess the ‘hot spots’ within a process that require more immediate attention. For process control, daily monitoring using ATP test kits will give you true total microbial quantity parameters to trend over time against process characteristics and performance.When utilizing ATP test kits it is important to remember that every process is different. During audits, relative comparisons from point to point are a reliable means to assess your process, while for daily monitoring it is important to establish a baseline trend before making control decisions.
NB: the QGO-M ™ method is approved at E 2694 for measurement of ATP in metalworking Fluids and is currently under ballot for approval as a means to measure ATP in fuels, fuel/water mixtures, and fuel-associated waters. And it has been extensively tested and validated for all types of fuels – conventional and non-conventional.
QGO-M is a trademark of LuminUltra Technologies Ltd (Canada). Kits are manufactured by LuminUltra.
AAQQQUUUAAA-TTTOOOOOOLLSSales of New Generation of ATP-metry kits for fast and true quantifi cation of microorganisms (Viable Culturable
& Viable but None culturable State in Bacteria)
The new generation of ATP-metry kits developped by LuminUltra (Canada) and commercialized by Aqua-tools is based on bioluminescence and allows measurements in less than 6 minutes on the fi eld for a higher reactivity for implementing preventive and corrective action.
The kits are implemented in the following types of installations:Cooling towers | Sanitary water | Production & distribution of potable water | Process water | Pure and ultra-
pure water | Biofi lm | Metalworking fl uids | Oil & Gaz | Cataphoresis baths | Surface treatment | Activated
sludge in water treatment | Bioreactors | Reuse water | Other… |
OTHER MARKETING TOOLS AVAILABLE
#1 Water testing Use of Quench-Gone™ Aqueous Kits as a tool to set up Risk Analysis and Routine Surveillance Programs for Sanitary Water and Cooling Tower Systems in Hospitals, Spa#2 ATP-metry technology for Autocontrol of Cooling Circuits
#3 Use of Total Control Biological (TCB™ / QG21 W ™) kits for operating the biological activity in Waste Water Treatment Plants#4 Use of Quench Gone Organic-Modifi ed™ kits to follow up the microbiological activity in the petroleum industry(« Upstream, Midstream and Downstream activities »)
QGO-M™ TEST PROTOCOL
QGO-M™ CALIBRATION
Put 2 drops of Luminase and 2 drops of UltraCheck 1 into an assay tube. Place immediately the assay tube in luminometer, hit “Enter” and record the RLU
UC1.
Perform only one calibration per sample set.If RLUUC1 < 5000, replace Luminase.
QQQGGGGOOO--MMMMM™™™™™ TTeeeeccchhnnnicccaaall DDDaaataa SSShheeeett
RLU
UltraCheck 1 Luminase
2 Drops (100μL)
Assay Tube
2 Drops (100μL)
For further information, refer to the instruction note #1
Remove
Attach
(1)
(2) 20mL Syringe
(3) Step 1 : Measure VolumeMix your sample. Remove the plunger from the 20ml syringe and attachthe fi lter. Pour the right volume (20mL of fuel or 5mL of bottom water)into the barrel of the syringe.
Push
Waste Vessel
20mL Syringe
(1)
(2)Step 2 : FiltrationSlowly push the sample through the fi lter into a waste receptacle, without pushing the plunger all the way down (ensure the fi lter remains wet). Detach fi rst the fi lter and then remove the plunger (always in this order).
LumiClean
5mL
Push
5 mL
Waste Vessel 20mL SyringeAttach
(2)
(3)
(1)
Step 3 : WashingRe-attach the fi lter to the syringe and add 5ml of LumiClean to the bar-rel. Slowly push the plunger to the bottom of the syringe barrel, forcing the LumiClean through the fi lter and into the waste receptacle. LumiClean is used to clean the fi lter and can be thrown up after this step.
(2)
(1)
(2)
(1)
Push
Waste Vessel20mL Syringe60mL Syringe
Step 4 : DryingDetach the fi lter and remove the plunger, and then attach the fi lter to a 60ml syringe. Dry the fi lter by pushing air through the fi lter.Same 60ml syringe can be used to dry fi lters up to 20 times.
QGO-M™ TEST PROTOCOL
1mL
UltraLyse 7
1 mL
Push
Extraction Tube
20mL SyringeAttach
(1)
(2)
(3)
9 mL
9mL
UltraLute Extraction Tube Diluted Extraction Tube
Assay Tube
Luminase
2 Drops (100μL)
RLU
100 μ
L
100μL
Diluted Extraction Tube Assay Tube
Step 5 : ExtractionRe-attach the fi lter to the 20ml syringe and add 1ml of UltraLyse 7. Slowly fi lter the UltraLyse 7 into a plastic tube.This extract can be stored refrigerated (2°-8°C) 7 days or 3 hours at 20°C.Make in serial the steps 1 to 5 for all the samples to be analyzed before the step 6.
Step 7 : AssayTransfer 100µl from the plastic tube to an assay tube and add 2 drops of Luminase. Immediately place the assay tube in luminometer, hit “Enter” and record RLU
cATP.
Step 6 : DilutionAdd 9ml of UltraLute into the plastic tube. Cap and mix. Immediately proceed to the next step!
QQGGGGOO--MMM™™™ VVoooluummmee ssaaammmppleee• Typical volume used for Fuel = 20mL• Typical volume used for Bottom Water = 5mL
QQGGGGOO--MMM™™™ iinttteerrprreeeetaaatiioon gguiiddellinnesss foor FFuel & WWaateer
For sales information, please contact Aqua-tools26, rue Charles Edouard Jeanneret 78300 Poissy - France Tel : + 33 (0)1 39 75 02 20Fax : + 33 (0)1 39 75 08 28 [email protected]
CALCULATION
Once calibration and tests performed, calculations must be done to obtain a proper ATP concentration in picograms per milliliter (pg/mL, 10-12g/mL):
emuloV (mL)SampleRLURLUAc (pg/mL)TP
UC
cATP 01 0001X=
QGO-M™ Technical Data Sheet
✔ !ACCEPTABLEFuel: < 10 pg ATP/ml
Water: < 100 pg ATP/ml
PREVENTIVE ACTION REQUIREDFuel: 10-100 pg ATP/ml
Water: 100-1000 pg ATP/ml
HEAVY CONTAMINATIONFuel: > 100 pg ATP/ml
Water: > 1000 pg ATP/ml
