the technologies of chemical metrology and quality

The Technologies of Chemical Metrology and

Quality Assurance of Measurement Results in

Food Analysis

Prof. Li Hongmei

National Institute of Metrology, China (NIM, China)

National Research Center of Certified Reference

Materials(NRCCRM)

Outline

1. Background

2. Metrological Traceability in Analytical Chemistry

Concept of metrological traceability

Key factors of measurement quality control

Method development

CRM production

Proficiency test

3. Establishment of the global measurement system in

mycotoxins

4. Summary

2

An inter-laboratory comparison held by EU in the year of 2000

Key factors for chemical

measurement

Measurement methods

Pre-preparation and reagent

Operator and environment

Measuring instrument

Reference material

Others

The measurement Results are accurate or not?

One time measurement

Globally effective

Primary Method

PRM

CRM

High Accuracy Method

Standard Method

analysis

SI unit

Chemical metrology system

Traceability: NMIs develop and establish the unbroken metrological traceability chain by providing the services of CRMs, PT and results assessment etc.

Comparability：The transverse connection of metrology, calibration and testing laboratory at different level

The Role of Chemical Metrology

Chemical Metrology vs. Free Trade

Test

Comparability

Traceability

One Test

Mutual Recognition

Global Recognition Free Trade

Sustainable Economic Development

Metrology Standard

Measurement Quality Assurance

specialized

production

Product

Consistency Product

Conformity

Precision Product

Quantification

Promoting trade Improving products

Sustainable Economic Development

product application

(market access)

product application

(consumer protection)

Conformity

Assessment

Reference Measurement Systems supports food safety

International Mutual

Recognition System:

International Organizations

Chemical

Metrology

System

CRM:

The key role

7

Grain food is a major nutrition source. Among 10 largest grain exporting countries in the world, 6 are

from APEC, such as the United states, Australia and Canada.

Grain safety main concerned with different contaminants, like mycotoxins, heavy metals,

pesticide residue, microorganism, etc.. Among them, mycotoxins and heavy metals are two

major harmful influence factors.

Grain safety is the basis of national economic security, and is the core of the production,

processing, trade, reserve and consumption balance.

Measurement and standards provide accurate and internationally recognized methods to assure

grain safety.

Harmonized global measurement and standards can improve quality control system in

developing countries, reduce economic loss and trade disputes.

The role of of measurement and standards in grain food

Outline

1. Background

2. Metrological Traceability in Analytical chemistry


Key factors in quality control of measurement

Method development

CRM production

Proficiency test


mycotoxins

4. Summary

9

VIM definition：

Property of a measurement result whereby the result can be related to a reference through a documented unbroken chain of calibrations, each contributing to the measurement uncertainty.

The term “Traceability” is replaced by “Metrological traceability”

a new definition as property of a measurement result which can be related to a reference

an evidence of measurands tracing to the primary standards which can realize the SI units

a documented unbroken chain of calibrations

2.1 Metrological Traceability

10

Metrological Traceability

ISO/IEC Guide 99:2007, “International vocabulary of metrology — Basic and general concepts and associated terms”

11

Unbroken chain of calibrations

Results

Calibration

Calibration

Calibration Results

Measured

SI /Reference

Measurement standard

Measurement procedure

Measurement unit

Measurements uncertainty

Measurements value &

Document

Document

Document

Outline

1. Background




Method development

CRM production

Proficiency test


mycotoxins

4. Summary

12

13

2.2 Key factors in quality control of measurement

Chemical metrology

Method

CRM Reference value for PT

Primary methods

High accuracy methods

Pure / solution CRM

Matrix CRM

Measurement standards and tools for metrological traceability

Calibration CRM1

Sample

Measurement result1

Higher level reference

Common reference

Trac

eab

ility

Matrix CRM1

Bias

Matrix CRM2

Bias

Higher level reference

Calibration CRM2

Measurement result2

Sample

comparability

comparability

14


Sample preparation techniques

Measurement characteristics

Measurement techniques

Advance material

Environment

Food safety

Medicine

Forensic science Security

Energy

Core capability principles in establishing chemical metrological traceability system

15


Outline

1. Background

2. Traceability control in chemical metrology

Concept of metrology traceability

Key factors in traceability control

Method development

CRM production

Proficiency test

3. Traceability system establishment in mycotoxin

4. Summary

16

17

Water content The related

structure

impurities

VOCs Non-volatiles

Qualitative

analysis by GC-

MS, LC-MS

Quantitative

analysis by GC, LC,

GCMS,LCMS

Karl Fischer

titrator

Verify by TGA

headspace GC-

MS / headspace

GC-FID

Verify by q-

NMR,TGA

Ash residue

method, TGA or

ICP-MS

Verify by TGA

Purity assessment of organic compound-Mass Balance Approach

(MBA)

P=1-WRS-WW-Wv-WNV

2.3 Method development

18

Procedure

Selection of internal

standard

Selection of solvent

Selection of

quantitative peaks

Optimization of

qNMR paramemeters

Determination

Potential primary method

Traceability can be established to SI units

Universal to almost organic compounds

Simple pretreatment, which may decrease

uncertainty

No identification of impurities of water,

solvent, inorganic…

One peak from analyte and one peak from

internal standard is enough

Advantage

QNMR

Chemical components measurement result and influence factors on metrological traceability

Method of matrix sample

Definition of

measurement

Sampling

Sample measurement

data processing and uncertainty evaluation

Result and application

Inaccurate or incomplete definition of the

measurement

Matrix effects

Instability of chemical composition ;

Complicated sample transformation

measurement process

Complicated uncertainty components and

combination method

19

Mea

sure

me

nt

cal

ibra

tio

n

Isotope Dilution Mass Spectrometry －－high accuracy method !!!

• Relative measurements, excellent precision and accuracy

compared with standard additions

• Ideal internal standard (the same compound) , Correction for

signal drift and matrix effects

• Quantitative separation processes are not required

• Once equilibration of the spike and analyte isotopes has

been achieved, the total recovery of the analyte is not

required

Isotope Dilution Mass Spectrometry

Optimizing chromatographic conditions

Preparation of stable isotope labeled

standard solution

Value

assignment

Matrix sample test

Ratios of analyte to isotopic internal standard

………

International

comparison

Matrix sample test

Yes

No

Optimizing mass spectrometry conditions

Yes No

CRM value assignment procedure by IDMS in organic targets

21

Matrix effect

sensitivity

selectivity

accuracy

Mass spectrum detection

Resolution

high throughput

high efficiency

Chromatographic Separation

Extraction recovery

Remove matrix

Interferences

concentration

Sample preparation

Key points of IDMS method

IDMS technique can effectively compensate the matrix effect in some

situation, matrix, instrumentation and other factors may still give rise to

variation of the peak area ratio of analyte to isotope and consequently lead to

unpleasant result.

22

Effect of composition of mobile phase and sample solvent on θ

40%

35%

30%

25%

0.80

0.85

0.90

0.95

1.00

1.05

30%

35%

40%

Th

e fa

cto

r o

f m

atr

ix e

ffec

t (θ

)

Ace

tonitr

ile in

disso

lutio

n sol

vent

Acetonitrile in mobile phase

When the content of acetonitrile in the mobile phase was increased, the

difference in ME of CAP and D5-CAP was getting smaller, or even disappeared. 23

Matrix effect -- Chromatographic Separation

Chloramphenicol in milk powder

Melamine 13C3-Melamine 15N3- Melamine 13C3-15N3-Melamine

N

N

N

NH2

H2N NH2

1 2

3 4

5

6 N

N

N

NH2

H2N NH2

13

13

13

15

1515

N

N

N

NH2

H2N NH2

13

13

13

N

N

N

NH2

H2N NH

2

15

1515

IS Increased mass Total mass The ratio( %)

13C3-melamine 3 39 7.69

13C3-15N3-melamine 6 84 7.14

15N3-melamine 3 45 6.67

ME of Melamine in Milk Powder

24

Matrix effect -- Mass spectrometry detection

Response Ratio of Melamine to Internal Standards

0.0 1.5 3.0 4.5 6.0 7.5 9.0

0.9

1.0

1.1

1.2

1.3

1.4

average:1.35

average:1.27

Th

e r

ati

o o

f th

e p

ea

k a

rea

The concentration of analyte，ug/kg

The ratio of peak area （melamine / 15

N3-melamine）

The ratio of peak area（melamine / 13

C3-

15N

3-melamine）

The ratio of peak area（melamine / 13

C3-melamine）

average:0.97

13C

13C-15N

15N

Different response ratio has been found in melamine to three internal standards

with same concentration, average values are 0.97, 1.27 and 1.35 respectively.

Matrix effect -- Mass spectrometry detection

Uncertainty evaluation

Instrumental

optimization

Matrix effect

Balance

linearity Balance

calibration

Sample weighing

CAP in milk powder

efficiency

weighing

SPE evaluation

separation

Pretreatment, chromatography separation, Sample solution

repeatability

Matrix removal enrichment Clean up

efficiency

Column evaluation

components matrix

extraction

separation

transfer

Method

parameter

Volumetric

flask

Standard

solution CAP dilution

Temperature D5-CAP dilution

MS condition

calibration

Calibration solution Uncertainty account for ：57%

Uncertainty account for :43%

Purity

matrix solvents

instrument

Linear

range

26

Chloramphenicol in milk powder

57%

Uncertainty evaluation--chloramphenicol in milk powder

27

Uncertainty evaluation

Outline

1. Background




Method development

CRM production

Proficiency test


mycotoxins

4. Summary

28

29

Value-Assignment of Calibrant

High pure substance

Identification analyte in calibrant

Purity /calibrant assignment method

Sample Analysis Competencies

Identification of analytes in sample

Extraction of analytes

Cleanup - separation of analytes

Transformation – conversion of analytes

Calibration approach for value-assignment of analytes in matrix

Verification method for value-assignment of analytes in sample

2.4 CRM production

Core competency

Food matrix CRMs in NIM

Till Sep. 2016，food related CRM: 451，pure & solution: 391；matrix: 60

0

2

4

6

8

10

12

14

16

18

20

Matrix type 19

15

12

9

7

5

3 3 2

0

5

10

15

20

25

30

35

40

45

Measurands

42

8 6

4 4 4 3 2 1 1

0

20

40

60

80

100

120

140

160

pesticide additive nutrient hormone packing contact

drug elements species

phthalate clinical

Pure CRM

137

77 68

36

20 20

8 8 6

30

Uncertainty

evaluation

Validation

Quantitation

Sample 1-chloramphenicol in milk powder

Chromatography

separation

Sample

preparation

MS

optimization

Xiu Qin Li, Qing He Zhang, Hong Mei Li，Analytica Chimica Acta, 807 (2014) 75– 83

Value（μg/kg） U（μg/kg）（k=2）

<0.004 --

0.915 0.067

31

Relative CRMs

CRMs No. Mass fractions U (%) k=2

purity of melamine GBW10058 99.5 % 0.5

melamine in milk powder

GBW10059 0.45 mg/kg 16

GBW10060 3.47mg/kg 14

GBW10061 16.5mg/kg 10

32

Outline

1. Background




Method development

CRM production

Proficiency test


mycotoxins

4. Summary

33

Specially designed inter-laboratory comparisons was

combined with external QC activities of field laboratories

and promote the establishment and spread of valid

national metrological traceability system.

Providing customized multi-level blind samples

Providing independent reference values

Providing technical analysis in aspects of reference materials used，method performance, measurement uncertainties and risk in

compliance assessment like false negative or positive results.

2.5 Proficiency test

34

PT on food safety organized by NIM

35

Year Name number of

participants

2010 Determination of chloropropanol in soy sauce 23

2011 Determination of benzo[a]pyrene in edible oil 25

2011 Determination of sodium thiocyanate in liquid milk 25

2012 Determination of lead and chromium in solution 31

2012 Determination of lead and chromium in milk powder 31

2012 Determination of lead and chromium in tea 31

2012 Determination of pigments in wine 130

2012 Determination of cadmium in rice 510

2013 Determination of aluminum in vermicelli 40

2013 Determination of 16 kinds of phthalate acid esters in Chinese spirit 15

2013 Determination of aflatoxin in liquid milk 15

2014 Determination of 16 kinds of phthalate acid esters in chinese spirit 37

2014 Determination of lead in infant powder 37

2014 Determination of aflatoxin M1 in milk 33

2015 Determination of clenbuterol in pork 8

2015 Determination of progesterone and methyltestosterone in milk 8

16 food safety related PTs cover 11 food matrix, about 13 kinds of measurands

Multi-targets and sample transfer means

•Performance of measurement methods

•Measurement capability of testing

laboratories

•Accuracy of the property value

of reference materials

•Preparation capabilities

•Certification capabilities

pilot laboratory

…

pilot laboratory

Lab1

Lab2

Lab3

Lab n

Lab1

Lab2 Lab3

Lab n

… 36

• Primay data analysis

•Regressed value for multi-point comparison

•Standard deviation estimate

•Performance evaluation

•Histogram

•Kernel density

•Normal distribution test

•Ordinary linear least square method → linear regression with errors in both axes

• Intercept ≠ 1 and slope ≠ 1 significantly:

• Big errors in the nominal property values of evaluated RMs

• Commutability

• Systematic error

•Standard deviation estimate of uncertainty weighed mean from expert

laboratories

•Standard deviation estimate of robust median from participants

•Shortest coverage interval

• Method performance

• Laboratory performance

• Analytical method dependence

• Matrix effect and commutability

• Technical advices

Integrated statistical and technical analysis

37

10

23

3

1

11

14

718

20

9

17

8

6

25

22

24

15

122

19

13

-6

-4

-2

0

2

4

-5 -4 -3 -2 -1 0 1 2 3 4 5

95%置信概率系列2 99%置信概率

0

20

40

60

0.9 1 1.1

Traceability control

Priority in selecting reference value

1. Measured or gravimetric prepared value by pilot laboratory

2. consensus value from collaborative expert laboratories

3. Consensus value from participants

4. Other methods

Tra

ce

ab

ilit

y

Ve

rifi

ca

tio

n

38

Priority in selecting measurement method

1) national primary apparatus or primary methods;

2) High accurate methods, the accuracy of them has been proved through the international measurement

comparisons;

3) reference laboratory system established through inter-laboratory study;

4) high precision methods using PRM or metrological traceable to provide measurement traceability.

CCQM-K24 Cadmium in rice

CCQM-P64,K56 Trace elements (calcium,

iron, zinc and copper) in whole fat soybean

powder

CCQM-K59 Nitrite in water

CCQM-K103 melamine in milk powder

CCQM- K106 Pb, As and Hg in cosmetic

material

CCQM- K89 Trace and Essential Elements in

Herba Ecliptae (As,Ca,Cd,Pb,Zn)…

Cd in rice with two levels of concentration

PT- nutritious elements in whole fat soybean powder

Quality evaluation of reference materials :nitrite in

water

Method performance assessment of rapid testing

methods for melamine in milk

PT- Hg in cosmetic material with two levels of

concentrations

PT- Cd,Pb,Cu in white atractylodes rhizome…

Median: 0.216 mg/kg

NIM value:

0.214mg/kg


International

comparison PT

PT International

comparison


39

GC-HRMS

LC-PDA

Raman Microscope

LC-MS-MS

奶粉5（16.3mg/kg) Z比分数

24 0610

6614 02

4965 71

42 64 51 7377 21

5552 7539 76 59 35 58 16 17 13 03 45 40 53 85 04 41 70 31 79 25 87 63 29 78 09 15 67 30 86 12 23 44 11 18 34 05 38 74 80 48 61 69 37 68

83 07 19 36 26 32 89 54 43 22 72 88 08 81 56 8250

84 28

-8.00

-7.00

-6.00

-5.00

-4.00

-3.00

-2.00

-1.00

0.00

1.00

2.00

实验室代码

Z比分数

｜Z｜< 2 82%

2<｜Z｜< 3 7.2%

｜ Z ｜＞ 3 10.8%

Melamine in milk powder

CCQM-K103 & P145

Melamine in liquid milk and milk powder 40

41

PT -- Aflatoxin M1 in milk

33 Labs 0.2 μg/kg

NIM prepared the PT sample and certified the reference value using

LC-IDMS

The certified value has good consistency with the medium value

26 of 33 labs got favorable results

Median Reference value

Outline

1. Background




Method development

CRM production

Proficiency test


mycotoxins

4. Summary

42

43

Laula Anfossi, Recent patents on food, nutrition & agriculture, 2010, 2,140-153

3 Establishment of the global measurement system in mycotoxins

44

3 Establishment of the global measurement system in mycotoxins

•Secondment

•Training

•PT

•Calibration solution

•Food matrix

•Calibration solution

•Matrix CRM

• Purity assessment

• High accuracy method

• Standard method Method

development CRM

Knowledge transfer, PT

Comparison

Calibrations:Aflatoxin B1,

Zearalenone, Patulin,

Deoxynivalenol,etc.

Matrix:Cereal, Dairy product,

apple juice, oil, nuts, meat

Collaboration with BIPM and other metrology institutes, to develop metrological

capacity and the metrology infrastructure in mycotoxin:

• To quantify mycotoxins in food accurately

• To support standardization in mycotoxin

• To realize world-wide free trade in food

Regional Metrology Organizations (RMOs)

APMP

NIM

ACRM

Polarity:

High Polarity(-2 4)

Low Polarity(-8 -2)

Molecular weight:

Low MW(0 ~ 300)

Medium MW(300 500)

High MW(500 800)

Priorities in Mycotoxins – Matrix CRM

Patulin

Zearalenone

Aflatoxin B1

Aflatoxin M1 Deoxynivalenol

Ochratoxin A

Fumonisim B

0

100

200

300

400

500

600

700

800

-6 -5 -4 -3 -2 -1 0 1 2 3

MW

PKow

MW vs. Polarity

Company Logo

Ochratoxin A in cereal etc.

Fumonisim B in maize etc.

Zearalenone in maize etc.

Deoxynivalenol in rice flour

Aflatoxin M1 in milk powder

Aflatoxin B1 in oil

Patulin in juice

Ochratoxin A in meat etc.

Aflatoxin B1 in peanut

Outline

1. General background




Method development

CRM production

Proficiency test


mycotoxins

4. Summary

47

1. Delivery service by CRMs and PT from NMIs are the important approach of

establishing metrology traceability system

2. The national measurement technical system is established through the

international mutual recognition to the core competency achieved by study on

features of analytical techniques, typical matrix and the characteristics of

analytes. It is the technical insurance of the formulating of standards and

laboratory accreditation.

3. High accuracy techniques of chemical metrology could evaluate the accuracy of

measurement results of testing as well as ensure their reliability and validity. It is

primary importance to the improvement of technical progress of advanced

measurement technologies in the field of food safety.

4. The establishment of Metrological Traceability system in Mycotoxin sustains

world-wide free trade.

4 Summary

Acknowledgement

Funded by the Ministry of Science and Technology of China

The technical group:

Dr. Li Xiaomin, Dr. Zhang Qinghe, Dr. Li Xiuqin,

Dr. Lu Xiaohua, Dr. Lu Hai, Mr. Guo Zhen, Dr. Huang Ting,

the technologies of chemical metrology and quality

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