Case Study: GlaxoSmithKline

Aim: To Determine the Best Method of Detection for Analysing

Nitrates in the Wastewater of a Large Pharmaceutical Industry.

Roisin Walsh

Laura Brophy

Contents

• Nitrates-Eutrophication• GlaxoSmithKline-Introduction to the

Pharmaceutical Company• Analytical Techniques• Environmental Protection Agency-IPC

Licensing and Monitoring• Discussion

Nitrates

The Pollution Effects Associated with the Emission of Nitrates

Nitrates

• Nitrate (NO3-) is a nutrient present in

fertilisers, as well as phosphate (PO4-).

• Excess amounts of nitrates and phosphates can enter rivers and streams through runoff from agricultural land and industrial and urban wastewater discharges, thus leading to eutrophication.

Eutrophication• When nutrient levels in a

waterbody are higher than natural levels, plant growth, especially algae, can occur. Algae remove oxygen from the water, which makes it difficult for other plants and animals to get enough oxygen to survive.

• This is known as eutrophication.

GlaxoSmithKline

An Introduction to the Pharmaceutical Company

GSK-Introduction• The Cork facility was

established in Carrigaline, in 1974, to manufacture bulk active pharmaceuticals.

• GSK employs a total of over 1,400 people in Ireland, including more than 400 at the Cork facility.

GSK-Development• GSK is the second largest pharmaceutical

company in the world. Some recent developments in the Cork facility include:1. 5 year - €225 million facilities investment programme,

which included a new state-of-the-art chemical development R&D pilot plant and laboratory.

2. An €8 million expansion of its chemical pilot plant and €7 million R&D investment to carry out research into the physical properties of active pharmaceutical ingredients and the reformulation of HIV products.

3. A €19 million investment for the development of nanomilling technology, made in late 2003.

GSK-Wastewater Discharge

• The GSK facility in Cork has its own wastewater treatment facility.

• It continuously monitors the wastewater being discharged.

• At present the method they use for nitrate detection is Cadmium Reduction with UV Detection. Ion Chromatography was tried, but found to be too slow.

Aims of Study

• The continuous expansion and development of the company will result in an increase in waste discharges.

• As a result, the most up to date techniques are required for the monitoring of all emissions from the plant.

• For the purposes of this case study, only the methods used for nitrate detection in wastewater will be assessed.

Methods of Nitrate Detection

Assessment of the Various Analytical Techniques Used to Analyse Nitrates in Wastewater

Methods of Nitrate Detection

• A number of analytical techniques used to detect nitrates will be discussed. These can be grouped under the following headings:

1. Electrochemical2. Capillary Electrophoresis3. Spectrometry4. Chromatography

1.Electrochemical Methods1. Voltammetric Methods:

– Lots of different electrodes are available including copper, nickel, cadmium and platinum.

– Using bare electrodes is not facile-kinetics of the charge transfer are slow, direct reduction of nitrate characterised by poor sensitivity and irreproducibility.

– By maintaining a large and highly active surface area the sensitivity can be increased. Can be done by simply introducing the appropriate metal salt. The analysis is relatively independent of the base electrode metal material, as the nitrate/nitrite reduction occurs at the freshly deposited metal layer.

1.Electrochemical Methods– By preparing the electrode ex situ, whereby the

electrode surface is conditioned in a plating solution of defined composition and then transferred to the analytical solution, greater control over the morphological features of the deposit is achieved.

– More recently electrode cleaning and activation have been achieved through the application of 20kHz ultrasound.

– Large number of assay type protocols, which utilize chemical properties such as the ability to nitrate aromatics and oxidise uranyl ions.

1.Electrochemical Methods

2. Potentiometric Methods: – Most common approach is the use of ISEs.

Detection limits for nitrate offer micromolar resolution coupled with good dynamic ranges spanning from 10-5 to 10-2M.

– Advantage-nitrate sensors can easily be coupled with continuous flow system or in flow system with sample injection such as FIA.

2.Capillary Electrophoresis

• Nitrate determination performed by UV detection at 214nm.

• Preparation stages may be required.

• Deproteinization can increase the lifetime of the system to 250-300 determinations.

2.Capillary Electrophoresis• Advantages:

1. Possibility of fast simultaneous detection of a wide variety of anions

2. Small sample requirements (nl)3. Low buffer consumption4. Instrumentation has automated sample loading,

requires little maintenance and is arguably more cost-effective

5. Use of capillary zone electrophoresis (CZE) has increased sensitivity x10 resulting in 0.1µM limit of detection

3.Spectrometry1. Greiss Assay Method:• Azo chromophore produced by

diazotisation of an aromatic amine by nitric acid.

• Absorption of the azo dye measured by VIS spec.

• Nitrate/nitrite levels are subsequently determined.

• Detection limit tends to be 0.02 - 2μM.

• It is possible to couple the Greiss assay to HPLC and FIA systems. This allows analysis of NO3- in complex matrices.

Common reaction pathways for the detection of NO3

-

3.Spectrometry2. Flow Injection Analysis:• FIA is a continuous flow

technique, suitable for rapid automated analysis of NO3

- in water.

• Reactive or detectable species of NO3

- produced by addition of reagents

• Reactive species can be sensed by a variety of flow-though detection devices.

• Possible to couple the FIA set-up to the Greiss assay method

FIA Detection System

FIA Sampling System

3.Spectrometry3. Fluorimetric

Determination:• Reagents convert NO3

- to detectable form.

• E.g. NO3- measured by the

oxidation of NO2- with

Ce(IV) which produces Ce(III).

• Presence of other redox species can interefere with detection of the Ce(III).

• Several disadvantges:a) Poor reproducibilityb) Toxic reagentsc) Time consuming procedured) Narrow detection range.

3.Spectrometry4. Gas-Phase

Chemiluminescence:• Conversion of NO3

- to NO2

-.

• Reduction of NO2- by KI

produces NO gas.• This gas reacts with O3 to

produce NO2*. This excited state of NO2

- emits IR chemiluminescence at ~ 600nm as it decays

• This detection system can give better detection limits than conventional UV-VIS

Chemiluminescence/GC system

3.Spectrometry5. Cadmium Reduction:• NO2

- in the sample is measured firstly.

• This is followed by reduction of NO3

- in the sample to NO2

- by passing it through a Cu/Cd glass column

• Subsequent measurement of the absorbance of the reduced solution is carried out spectrophotometrically.

• Comments:– Requires specialized glass

columns for accurate analysis.

– Technique involves handling and disposal of toxic Cadmium and hazardous phenol.

– Method is subject to interference from other ions naturally present in water.

• Method most often used for NO3

- analysis.

4.Chromatography1. HPLC:• Not regularly used by

environmental laboratories for NO3

- analysis.• HPLC analysis of NO3

- can achieve detection range of 0.05 mg/l – 300 mg/l

• Separation is carried out by HPLC - detection often carried out with UV spec.

• HPLC can be coupled to the Greiss assay protocol

• No modification of analyte• Speedy analysis

• Systems can be automated – leading to lower running costs.

Typical HPLC set-up

4.Chromatography2. Ion Chromatography:• EPA Method 300 is the US EPA-

approved method for analysis of NO3

- - involves the use of Ion Chromatography.

• UV detection is usually carried out after the sample elutes from the column.

• No toxic/hazardous materials used/generated.

• Simultaneous detection of a number of anions in the sample

• Direct sample introduction – no derivatisation required

• Sample pre-treatment – usually filtration – required.

• Sensitive method, low LOD

• Expensive.• Instruments require frequent

maintenance.• Time-consuming and expensive

Typical Ion Chromatography system set-up

EPA and EMAS

Their Role in Monitoring the Environmental Impacts from a

Large Pharmaceutical Company

EPA

• The aim of the EPA is to – To protect and improve the natural

environment for present and future generations, taking into account the environmental, social and economic principles of sustainable development.

• One aspect of this is the monitoring of industries and any emissions and effluents coming from them through a licensing system.

IPC Licensing• The EPA are in charge of the

Integrated Pollution Control (IPC) licensing in Ireland.

• The EPA carry out visits to IPC licence holders, in most cases to monitor emissions. These visits are usually unannounced. Inspections are carried out both before and after an IPC licence has been granted.

• It is also required that the licensee monitors and reports on the environmental performance of their facility, with reports being submitted on a regular basis. An Annual Environmental Report is also submitted to the EPA.

Urban Waste Water Discharges in Ireland-EPA Report 2000-2001

• Each activity that requires a licence is categorised by the EPA.

• GSK is categorised in the Chemicals section, as 5.6-The manufacture of pesticides, pharmaceuticals or veterinary products and their intermediates.

Urban Waste Water Treatment Regulations, 2001

• According to this legislation, the minimum percentage of reduction for total nitrogen is 70-80%.

• The concentration varies depending on the population equivalent:– 15mg/L 10,000-100,000p.e.– 10mg/L >100,000p.e.

• The reference method used for measurement is molecular absorption spectrophotometry.

Auditing-EMAS• The EU Eco-Management and Audit Scheme

(EMAS) is a management tool open to all economic sectors including public and private services to evaluate, report and improve their environmental performance.

• Participation is voluntary and extends to public or private organisations operating in the European Union and the European Economic Area (EEA).

• The Irish National Accreditation Board is the competent body in Ireland for the administration of the EMAS scheme.

• Its aim is to recognise and reward those organisations that go beyond minimum legal compliance and continuously improve their environmental performance.

Discussion

Summary and Conclusions of Findings

Methods

• Overall the most widely used nitrate analysis method is Cadmium Reduction.

• However, this involves the use of toxic chemicals and as such the recommendation of this case study is that Ion Chromatography be used by large pharmaceutical companies for nitrate analysis.

• By coupling IC with FIA, the rate and efficiency of analysis is improved.

Legislation

• The method used by GSK for nitrate analysis needs to be approved by the EPA, and should provide results comparable with the nitrate targets as stated in the companies IPC Licence.

• The method should also be comparable to the reference nitrate analysis method (molecular absorption spectrometry), as set down by the EPA.

References• Detection and Determination of Nitrate and

Nitrite: A Review, M. J. Moorcroft, J. Davis, R. G. Compton, 2001. Talanta, 54, 785-803.

• Optimising the Determination of Nitrate and Phosphate in Sea Water with Ion Chromatography Using Experimental Design, I. Dahllof, O. Svensson, C. Torstensson, 1997. Journal of Chromatography, 771, 163-168.

• www.epa.ie• http://www.gsk.com/ehs/pdf_excel/ehs.pdf