2011-07-1_Adulteration Food Chem Uni Lecture_BRA Cm

8/3/2019 2011-07-1_Adulteration Food Chem Uni Lecture_BRA Cm

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Markus Lipp, Ph.D.Director, Food Ingredient Standards

US Pharmacopeia, Rockville, [email protected]

Food Quality, Safety, and the role ofthe Compendial Standards

Ice Breaker: Hypothetical scenario

• You are the chief chemist in charge of QA/QCand scientific affairs for a bread company

• Reports from customer hotline

– Nausea and vomiting after wheat breadconsumption

– Ammonia odor

• Determined that wheat flour used in yourproduct is likely the issue

Wheat flour certificate of analysis

DESCRIPTION OFGOODS:

WHEAT FLOUR

PACKING: PACKED IN 25KGBAGS

ORIGIN: USA

TEST SPEC RESULTMoisture NMT 14.0% 7.5%

Ash ( On drybasis)

NMT 2.0% 0.89%

Protein (On dry

basis N x 5.7):

NLT 9.0% 9.1%

Vomitoxin NMT 1 ppm < 18.5 ppb

Vomitoxin (Deoxynivalenol)?

Result:

< LOD (18.5 ppb)

ELISA



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What is happening here?

Test WG:Why?

9.0%

Control flour 9.0% 9.0%

Test flour 9.0% 8.0 %

Wheat flour specsfor protein content

NIR spectral analysis of wheat flour

Courtesy of K. Norris, 2009

Wavelength used for

protein measurement

due to N-H vibrations

(2180 nm = 4587 cm–1)

Hold that thought!

• Understand:

– The role of food chemistry and compendialstandards in food quality and food safety for foodingredients and additives

– The importance of methods of analysis andreference standards for food ingredient analysis

• Apply knowledge:

– As future food science professionals

Lecture objectives



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Food quality, safety, and chemistry

Food Quality Food Safety

– Physical hazards

– Microbiological hazards

– Chemical hazards

Organoleptic

properties

Toxicological

properties


• Is the quality of an incoming ingredient goodenough for its intended use – “food-grade” ?

• How do you know that is safe (toxicologically)unless you know something about itscomposition (chemical)?

• If the ingredient is safe, how do you know thatthe material you purchased is chemically thesame as the material used to carry out the

safety evaluation?


• How do you know if the ingredient has anunacceptable level of a contaminant(unintentional impurity inherent to raw

material or formed during processing)? – Examples: Metal impurities, organic

solvents

Certificate of analysis (CoA)

• From buyer: Procurement specifications

• From seller: Certificate of analysis

• What test methods were used? Are themethods reliable? Validated?

• What reference standards were used?Are the standards reliable?



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Certificate of analysis (CoA) How to measure is important

• Analyzing for the same constituentor property using different methodscan yield different results

Examples to demonstrate importance of…

• Method validation: assessingperformance characteristics for analyticalmethods

– Specificity/Selectivity: Ability to distinguishanalyte from other similar components inmatrix

– Accuracy: Closeness to true value

– Limit of detection: Lowest amount ofanalyte that can be detected

– 5 other parameters

• Reference standards

Example 1: Measuring water content (specificity)

• Water content measurement

– Same sample analyzed for water content by “Losson Drying” and “Karl Fisher” methods

– Data: 0.088% vs 0.068%, respectively

• Loss on drying• Principle: Evaporation of volatile compounds

• Only water evaporating?

• LOD results 30% higher than KF, why?

• Results highly dependent upon conditions used

• Karl Fisher• Principle: Reduction of iodine by SO2 in the presence

of H2O, titration



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Example 1: Measuring water content (specificity)

• Why is water important?

– Water activity <-> foodsafety/quality

– Diluting a product with water is aneasy way to economicallyadulterate the product

• Methodology, Pre-1996 (FCC IV) – Hydrogen sulfide method (colormetric test)

• Metallic impurities colored as sulfides

• Visual comparison with known leadstandards

• ISSUES: – Not sensitive enough to work with limits below

10 ppm

– Not very accurate (next slide)

– Not very selective

Example 2: Measuring Pb

Example 2: Measuring Pb impurities (accuracy)

0

20

40

60

80

100

120

Pb As Se Sn Sb Cd Pd Pt Ag Bi Mo Ru In Hg

Elements

A v e r a g e % R e c o v e r i e s

USP Results

ICP-MS Results

Hydrogen Sulfide

Method Results

Lewen, N. et al J. Pharm. & Biomed. Anal. 35 (2004) 739-752

Example 3: Measuring metal impurities (limit

of detection)

• Specification requires “less than 5 ppm lead”

• CoA result: ND

• What does this mean?

– Less than the limit of detection of the method

• What if limit of detection for method is greaterthan 5 ppm?

– Actual concentration could be more than 5 ppm!



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Example 4: Reference standards:

the phoenician balance Example 4: Reference standards

• 1) Qualitative reference standard

– To substantiate the identity/ authenticity of an ingredient you cancompare to an authentic referencestandard

• 2) Quantitative reference standards

– Chemical “yard stick”

Qualitative reference standards

IR spectrum from authentic

glycerin

IR spectrum from test sample

(diethylene glycol)

Quantitative reference standards

HPLC chromatogram from stevia extract

Rebaudioside A

•How to determineconcentration?

Need an external

reference

standard!



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Standardization challenge!• Who sets the minimum standard?

– Who determines what minimum qualityspecifications (purity, impurities) are appropriate forfood ingredients?

– How do you ensure they are set in an objective andfair manner?

• How do you ensure that everyone’ s analyticalresults are comparable?

– Who sets the methods? Who validates testprocedures?

– How do you ensure that everyone is using thesame reference standards?

Food ingredients compendial

standards• What is a compendium?

– Wikipedia: “A compendium is a concise, yetcomprehensive compilation of a body ofknowledge”

• In the case of food ingredient standards,a compilation of chemical test methodsand specifications to establish quality andthus safety of individual ingredients.

– Comprehensive analysis of ingredients

(Identity, Purity, Impurities)

• Is this lot of material what the food ingredientseller claims it to be? How do you test for this?

– IDENTITY

• How pure is it? What purity is acceptable for

“food-grade” ? How do you test for this? – PURITY

• What kind of impurities does it contain? Whichones are important and what levels areacceptable? How do you test for these?

– IMPURITIES

Food ingredients compendial

standards Examples• The Food Chemicals Codex (FCC )

• The United States Pharmacopeia

• The Joint FAO/WHO Expert Committee on FoodAdditives (JECFA’ s)

“Combined Compendium of Food AdditiveSpecifications”

• Codex Alimentarius

• Other compendia



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What is the Food Chemicals Codex (FCC )?

• FCC is a compendium that establishes

internationally recognized scientific standards

to promote uniformity of quality and added

assurance of safety for food ingredients.

• FCC standards set using an objective, science

evidence–based, transparent process

• Food Quality↔ Safety

– How do you know it is safe if you don’t know whatit is? (identity and purity)

• Methods and limits for contaminants (inherentor process related impurites)

• First line of defense against economicadulteration

• Validated methods

• Developed by independent, volunteer scientificexperts through a transparent process

The need for FCC ?

What do FCC standards look like?

• Monographs

• Specifications for identity

(authenticity), purity and impurities

• Supporting test procedures and

reference standards



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Scientific utility of FCC

• Students

– Source, method of manufacturer, physicochemical

characteristics, CAS, INS, FEMA, and function(s) of

food ingredients

– Methods of analysis

• Validated analytical methods for specific food

ingredients

• Validated analytical methods in Appendices

– Potential impurities and contaminants in food

ingredients

– Validated analytical methods

Scientific utility of FCC

• Industry / Regulatory

– Use FCC specifications, supporting test procedures,

and associated reference materials to establish the

food-grade quality and safety of their ingredients,

and thus a stronger marketing presence

– Ensure that food ingredient in commerce is

equivalent to that which was toxicologically evaluated

– Aid to effective self-regulation by industry

• Specifications often referenced in agreements for sale

and receipt of food ingredients

– Source of food ingredient GMPs

The future of food chemistry to support compendial

testing standards

• Economically motivated adulteration

• Authentication and characterization of

botanically derived, complex mixture food

ingredients

• Nanotechnology

• Functional foods and nutraceuticals

• Green technology and sustainability

Definition of Economically Motivated Adulteration

“The fraudulent addition of non-authentic

substances or removal or replacement of

authentic substances without the purchaser’s

knowledge for economic gain of the seller”

Also known as: “Food Fraud”, “Food Counterfeiting”, “Intentional

Adulteration”



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Recipe for EMA

• Rising raw material prices

• Complex supply chains

• Complex and variable

compositions

• Sophisticated fraudsters

• Lack of tools to prevent

• Accepting culture

Significant

Fraud

opportunity

Sophisticated Fraudsters and Creative Fraud

Fraudster “designs”adulterant to evade existingQA system

QA system reacts bydeveloping new tests

Public health threat of EMA

• Food safety assumes a known composition

• Next adulterant is unknown

Food safety

collapses to a

singularity

The fraudster’s ethics &

knowledge define food

safety throughout the

whole supply chain

Reducing the Risk of EMA: Multi-Pronged Approach

Testing Supply Chain

Management

• Know your suppliers, andtheir supply chains

• Supplier qualifications and

audits

• QA systems

• GMPs

• HACCP

• “Anchors” food safetysystems in reality by

guaranteeing ingredient

authenticity

• Quality systems built on

this assumption



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(1) Test for the absence of a known

adulterant

Two Analytical Approaches To Verify Ingredient Integrity

Disadvantages:

– Known does not guarantee detection

2007 melamine in wheat gluten & 2008

melamine in milk

– Cannot feasibly test for all known adulterants

– Cannot detect unknown adulterants

(2) Test for what is supposed to be there

(compendial approach)

– Detect EMA by:

• Decrease in puritytest result

• Abnormal identity test result

Two Analytical Approaches To Verify Ingredient Integrity

Advantage:

– Can detect both known and unknown

adulterants when present at EMA levels

State of Compendial Testing Standards

• Identification Tests

– Virtually non-existent for many widely used complex food

ingredients (e.g. wheat gluten, whey)

– Many outdated non-specific wet-chemistry tests that are

not sufficient to detect and deter EMA• Flame tests, solubility tests

• Assay (Purity) Tests

– Many outdated non-specific wet-chemistry tests that are

not sufficient to detect and deter EMA

• Indirect purity methods (Kjeldahl for total protein)

• Titrimetric methods

• The technology exists, but…

• R&D is necessary to adapt it into viable

compendial tools to detect and deter EMA

Moving Testing Technology into the 21st Century



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Opportunities to Advance Compendial Testing Standards

• Higher selectivity, instrumental methods

– Identification Tests

• Non-targeted analysis using IR, Raman, or NMR + chemometrics

• Semi-targeted (chemical fingerprinting) using LC-UV/Vis, GC-FID,

LC-HRMS, GC-MS, CE + chemometrics

• Isotope ratio fingerprinting: IRMS, SNIF-NMR

• Other: PCR; DSC; Rheological Methods; Microscopy

– Assay (Purity Tests)

• Instrument LC or GC methods

Back to wheat flour!

What Actually Happened?

• 1980’ s: Urea added to wheat to artificiallyincrease the nitrogen content -> apparentincrease in protein

• Where? – Minnesota, USA

Folkenberg J, Nelson R, Snider S. 1990. Spiked wheat - Schuler Grain Co. uses additive to artificially boost the protein

content of its wheat. FDA Consumer 24 (Dec.)

Why urea?

• Classic case of economically motivatedadulteration: “The fraudulent addition of non-authentic substances or removal or replacement ofauthentic substances for economic gain without the

knowledge of the purchaser.”

• Urea falsely measured as protein by twointernationally recognized protein assays

– Nitrogen-based and NIR total protein



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Economics*

Wheat commodity pricing based on protein content

Premium wheat (12% P) = $13.12 / bushel

Regular wheat (11.5% P) = $12.12 / bushel

Recipe for “Fake Premium Wheat” with 0.06% urea:

100 lbs regular wheat (11.5% P) + 1 lb water + 3 ouncesurea

Cost of “Fake Premium Wheat” = $12.02 / Bushel

Profit = $1.10/bushel = $880 / truckload

*DeVries, 2009. www.usp.org/goto/proteins

Chemistry

• 46.7% nitrogen

• Kjeldahl response = Protein

• Dumas (combustion) response = Protein

• NIR response = Protein – WHY????

Chemistry of NIR

Absorption bands are weak overtones orcombinations of fundamental stretchingvibrational bands that occur in the mid-infraredregion

Chemistry of NIR

• NIR Diffuse-reflectance used for grain proteinanalysis, calibrated against Kjeldahl/Dumas

– NIR does not sense nitrogen, but rather

overtones of NH vibrations of the amino acids tosense protein

– Bands at 2055, 2100, and2180 nm arecommonly used for protein

– Urea has absorption bands at2197 and 2162 nmcorresponding to NH vibrations

McDonald CE, Bruns . 1988. Cereal Food World 33(4):367-9.



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NIR spectral analysis of wheat flour


Wavelength used for

protein measurement


(2180 nm = 4587 cm–1)

NIR spectrum of urea


Wavelength used for

protein measurement


(2180 nm = 4587 cm–1)


Importance to food chemistry?

• Incidents such as urea and melamine

adulteration have underscored the

importance of having reliable routine QA

methods that measure with a high degree

of selectivity/specificity

• Ensuring the safety of the food supply



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Moving forward for food protein analysis

• Authentication methods to detect

abnormalities/aberrations resulting

from significant level of adulteration

• New quantitative methods capable of

selectively and precisely determining

total protein contents

– To replace total nitrogen (Kjeldahl and

Dumas) approaches

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Deliberately adulterated milk by FTIR and PCA

-0.5 -0.4 -0.3 -0.2 -0.1 0 0.1 0.2 0.3 0.4

-0.1

-0.05

0

0.05

0.1

0.15

Score PC2

S c o r e

P C 5

Melamine+water

Cyanuric acid

Urea Melamine

Ammonium sulphate

• Normal milk• Adulterated milk

*Juan Romero, Wisconsin Dairy Center

Conclusions• Specifications play an important role in the quality

and safety of food ingredients

• The method of analysis and reference standard

used to assess conformance to a specification can

influence the outcome

• FCC is a source of minimum quality specifications

for the identity, purity, and impurities of food

ingredients along with supporting validated test

procedures

• FCC standards benefit industry, regulators, and

consumers, and promote the quality and safety of

food ingredients

Conclusions• There are many critical needs in the area of

compendial testing standards that will rely on

advances in food chemistry

• There are many “at-risk” ingredients that will

require substantial efforts to establish reliablestandards to prevent food fraud

• Food and agricultural science should be driving

these multi-disciplinary research efforts



Questions?

2011-07-1_Adulteration Food Chem Uni Lecture_BRA Cm

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