HPLC for Food Analysis Agilent

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Copyright Agilent Technologies Company 1996 2001, All rights reserved Reproduction adaption or translation. without prior written permission is prohibited except as. allowed under the copyright laws,Printed in Germany. www agilent com chem September 01 2001,Publication Number 5988 3294EN. The fundamentals of an,alternative approach to,solving tomorrow s. measurement,challenges,Gratzfeld H sgen and,Rainer Schuster.
Acknowledgements,We would like to thank Christine,Miller and John Jaskowiak for. their contributions to this primer,Mrs Miller is an application. chemist with Agilent Technologies,and is responsible for the. material contained in chapter 5,Mr Jaskowiak who wrote chapter 7. is a product manager for liquid,chromatography products at.
Agilent Technologies,Copyright Agilent Technologies Company. 1996 2001 All rights reserved Reproduction,adaption or translation without prior. written permission is prohibited except,as allowed under the copyright laws. Printed in Germany September 1 2001,Publication Number 5988 3294EN. Preface Modern agriculture and food processing often involve the. use of chemicals Some of these chemicals and their func. tions are listed below, Fertilizers increase production of agricultural plants.
Pesticides protect crops against weeds and pests, Antibiotics prevent bacteria growth in animals during. Hormones accelerate animal growth, Colorants increase acceptability and appeal of food. Preservatives and antioxidants extend product life. Natural and artificial sweeteners and flavors improve. the taste of food, Natural and synthetic vitamins increase the nutritive. value of food,Carbohydrates act as food binders, Such chemicals improve productivity and thus increase. competitiveness and profit margins However if the, amounts consumed exceed certain limits some of these.
chemicals may prove harmful to humans, Most countries therefore have established official tolerance. levels for chemical additives residues and contaminants in. food products These regulations must be monitored care. fully to ensure that the additives do not exceed the pre. scribed levels To ensure compliance with these regulatory. requirements analytical methods have been developed to. determine the nature and concentration of chemicals in. food products Monitoring of foodstuffs includes a check. of both the raw materials and the end product To protect. consumers public control agencies also analyze selected. food samples, High performance liquid chromatography HPLC is used. increasingly in the analysis of food samples to separate and. detect additives and contaminants This method breaks. down complex mixtures into individual compounds which. in turn are identified and quantified by suitable detectors. and data handling systems Because separation and detec. tion occur at or slightly above ambient temperature this. method is ideally suited for compounds of limited thermal. stability The ability to inject large sample amounts up to. 1 2 ml per injection makes HPLC a very sensitive analysis. technique HPLC and the nondestructive detection tech. niques also enable the collection of fractions for further. analysis In addition modern sample preparation tech. niques such as solid phase extraction and supercritical fluid. extraction SFE permit high sensitivity HPLC analysis in. the ppt parts per trillion range The different detection. techniques enable not only highly sensitive but also highly. selective analysis of compounds,Hydrophilic HPLC,Amino acids Inorganic ions. Volatile Synthetic Sugars,carboxylic,acids Glyphosate food dyes Sugar. Aldehydes Enzymes,Ketones PG OG DG,Sulfonamides Glycols phenols Aflatoxins.
Fatty acids Antibiotics,Nitriles BHT BHA THBQ,Polarity antioxidants Flavonoids. Nitrosamine,Organo Alcohol,PAHs Anabolica Natural food dyes. phosphorous, TMS pesticides Aromatic amines Fat soluble vitamins. derivative PCB, Essential oils Polymer monomers Triglycerides Phospho lipids. C2 C6 hydrocarbons Fatty acid Aromatic esters,methylester.
Hydrophobic,Volatile Volatility Nonvolatile, Match of analyte characteristics to carrier medium. Its selective detectors together with its ability to connect a. mass spectrometer MS for peak identification make gas. chromatography GC the most popular chromatographic. HPLC separates and detects at ambient temperatures For. this reason agencies such as the U S Food and Drug. Administration FDA have adopted and recommended, HPLC for the analysis of thermally labile nonvolatile highly. polar compounds, Capillary electrophoresis CE is a relatively new but rap. idly growing separation technique It is not yet used in the. routine analysis of food however Originally CE was applied. primarily in the analysis of biological macromolecules but. it also has been used to separate amino acids chiral drugs. vitamins pesticides inorganic ions organic acids dyes and. surfactants 1 2 3, Part 1 is a catalog of analyses of compounds in foods Each. section features individual chromatograms and suggests. appropriate HPLC equipment In addition we list chromato. graphic parameters as well as the performance characteris. tics that you can expect using the methods shown In part 2. we examine sample preparation and explain the principles. behind the operation of each part of an HPLC system sam. pling systems pumps and detectors as well as instrument. control and data evaluation stations In the last of 11 chap. ters we discuss the performance criteria for HPLC which. are critical for obtaining reliable and accurate results Part 3. contains a bibliography and an index, Part One Chapter 1 Analytical examples of food additives.
The HPLC Approach Acidulants 2,Antioxidants 4,Preservatives 6. Artificial sweeteners 8,Colorants 10,Flavors 12,Vanillin 12. Bitter compounds hesperidin and naringenin 14,Chapter 2 Analytical examples of residues and. contaminants, Residues of chemotherapeutics and antiparasitic drugs 16. Tetracyclines 18,Fumonisins 19,Mycotoxins 21,Bisphenol A diglydidyl ether BADGE 24.
Pesticides 26,Carbamates 28,Glyphosate 29,Chapter 3 Analytical examples of natural. components,Inorganic anions 32,Triglycerides and hydroperoxides in oils 35. Triglycerides in olive oil 37,Fatty acids 38,Carbohydrates 40. Vitamins 42,Water soluble vitamins 42,Fat soluble vitamins 45. Analysis of tocopherols on normal phase column 46,Biogenic amines 48.
Amino acids 50,Peptides 52,Part Two Chapter 4 Separation in the liquid phase. The Equipment Basics Separation mechanisms 58,Reversed phase materials 58. Ion exchange materials 58,Size exclusion gels 59,Adsorption media 59. The advent of narrow bore columns 59,Influence of column temperature on separation 60. Chapter 5 Sample preparation,Sample preparation steps 62.
Automation 62,Ultrasonic bath liquid extraction 63. Steam distillation 64,Supercritical fluid extraction 64. Liquids 65,Liquid liquid extraction 65,Solid phase extraction 65. Gel permeation chromatography 66,Guard columns 67,Chapter 6 Injection techniques. Characteristics of a good sample introduction device 70. Manual injectors 71,Automated injectors 72, Autosampler with sample pretreatment capabilities 72.
Derivatization 73,Chapter 7 Mobile phase pumps and degassers. Characteristics of a modern HPLC pump 76,Flow ranges 76. Gradient elution 76,Gradient formation at high pressure 77. Gradient formation at low pressure 77,Pump designs for gradient operation 78. Low pressure gradient Agilent 1100 Series pump 78, High pressure gradient Agilent 1100 Series pump 80.
Degassing 82,Helium degassing 83,Vacuum degassing 84. Chapter 8 Detectors,Analytical parameters 87,Limit of detection and limit of quantification 87. Selectivity 87,Linearity 88,Qualitative information 88. UV detectors 89,Diode array detectors 90,Three dimensions of data 91. Fluorescence detectors 95,Cut off filter 96,Signal spectral mode 96.
Online spectral measurements and,multi signal acquisition 96. Multisignal 97,Electrochemical detectors 98,Electrode materials 99. Flow cell aspects 99,Automation features 100,Mass spectrometers 101. API interfaces 102,Refractive index detectors 104,Chapter 9 Derivatization chemistries. Addition of UV visible chromophores 108,Addition of a fluorescent tag 109.
Precolumn or postcolumn 109,Automatic derivatization 110. Chapter 10 Data collection and evaluation techniques. Strip chart recorders 112,Integrators 113,Personal computers 114. Local area networks 117,Networked data systems 118. Chapter 11 Factors that determine performance in HPLC. Limit of detection and limit of quantification 121. Accuracy and precision 122,Qualitative information 123. Part Three References 125,References and Index,A demonstration.
of liquid chromatographic,separations in,food analysis. Analytical examples,of food additives, Acidulants Sorbic acid and citric acids are commonly used as. acidulants4 and or as preservatives Acetic propionic. succinic adipic lactic fumaric malic tartaric and, phosphoric acids can serve as acidulants as well Acidulants. are used for various purposes in modern food processing. For example citric acid adds a fresh acidic flavor whereas. succinic acid gives food a more salty bitter taste In. addition to rendering foods more palatable and stimulating. acidulants act as, flavoring agents to intensify certain tastes and mask. undesirable aftertastes,buffering agents to control the pH during food.
processing and of the finished products,preservatives to prevent growth of microorganisms. synergists to antioxidants to prevent rancidity and. viscosity modifiers in baked goods, melting modifiers in cheese spreads and hard candy. meat curing agents to enhance color and flavor,Sample preparation. Sample preparation depends strongly on the matrix to be. analyzed but in general steam distillation and solid phase. extraction techniques can be used,Chromatographic conditions. High performance liquid chromatography HPLC with,UV visible diode array detection UV DAD has been.
Isocratic Detector applied in the analysis of citric acid in wine and in a vodka. pump Auto Column VWD DAD, vacuum sampler compart or refractive mixed drink Retention time and spectral data were used as. degasser ment index identification tools,Control and. data evaluation,Sample preparation filtration,Column 300 x 7 8 mm BioRad mAU. 1 Oxalic acid 7 Lactic acid,HPX 87 H 9 m 400,2 Citric acid 8 Glycerol. Mobile phase 0 0035 M H2SO4 3 Tartaric acid 9 DEG,isocratic 300 1.
4 Malic acid 10 Acetic acid,Flow rate 0 6 ml min,5 Sulfur trioxide 11 Methanol. Column compartment 65 C 6 Succinic acid 12 Ethanol. Injection volume 10 l,Detector UV VWD,detection wavelength. White wine,192 nm or 210 nm 2 4 6 7 8 9 10 11 12,0 5 10 15 20 25. Analysis of acidulants in white wine,Conditions as above except. Citric acid,Mobile phase 0 007 M H2SO4 mAU Glucose 20.
Sample spectrum,isocratic 100 Fructose,Citric acid overlaid with. Detector UV DAD library spectrum,190 Wavelength nm 276. HPLC method performance,Limit of detection 100 ng injected amount. S N 2 equivalent to,2 ppm with 50 l,injected volume. Repeatability of 0,0 5 10 15 20,RT over 10 runs 0 1.
areas over 10 runs 3,Analysis of citric acid in vodka. 4 Official Methods of Analysis Food Compositions Additives Natural. Contaminants 15th ed AOAC Arlington VA 1990 Vol 2 Official Method. AOAC 986 13 quinic malic citric acid in cranberry juice cocktail and. HPLC for Food Analysis The fundamentals of an alternative approach to solving tomorrow s measurement challenges Angelika Gratzfeld H sgen and Rainer Schuster A Primer Acknowledgements We would like to thank Christine Miller and John Jaskowiak for their contributions to this primer Mrs Miller is an application chemist with Agilent Technologies and is responsible for the material contained

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