GB 5009.208-2016_English: PDF (GB5009.208-2016)
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Food safety national standard -- Determination of biogenic amines in food
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GB 5009.208-2016
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| GB/T 5009.208-2016 | English | 219 |
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Determination of biogenic amine in foods
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GB/T 5009.208-2016
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| GB/T 5009.208-2008 | English | 279 |
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Determination of biogenic amine in foods
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GB/T 5009.208-2008
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| Standard ID | GB 5009.208-2016 (GB5009.208-2016) | | Description (Translated English) | Food safety national standard -- Determination of biogenic amines in food | | Sector / Industry | National Standard | | Classification of Chinese Standard | X09 | | Word Count Estimation | 12,142 | | Date of Issue | 2016-12-23 | | Date of Implementation | 2017-06-23 | | Older Standard (superseded by this standard) | GB/T 20768-2006; GB/T 5009.208-2008; GB/T 5009.45-2003 Partial; SN/T 2209-2008 | | Regulation (derived from) | National Health and Family Planning Commission Notice No. 17 of 2016 | | Standard ID | GB/T 5009.208-2016 (GB/T5009.208-2016) | | Description (Translated English) | Determination of biogenic amine in foods | | Sector / Industry | National Standard (Recommended) | | Word Count Estimation | 11,176 | | Standard ID | GB/T 5009.208-2008 (GB/T5009.208-2008) | | Description (Translated English) | Determination of biogenic amine in foods | | Sector / Industry | National Standard (Recommended) | | Classification of Chinese Standard | C53 | | Classification of International Standard | 67.040 | | Word Count Estimation | 7,743 | | Date of Issue | 2008-11-21 | | Date of Implementation | 2009-03-01 | | Drafting Organization | China Disease Prevention and Control Center of Nutrition and Food Safety | | Administrative Organization | Ministry of Health | | Regulation (derived from) | National Standard Approval Announcement 2008 No.22 (Total No.135) | | Proposing organization | People's Republic of China Ministry of Health | | Issuing agency(ies) | Ministry of Health of People's Republic of China; Standardization Administration of China | | Summary | This standard specifies the foods serotonin, spermidine and spermine content determination, ��-phenylethylamine, putrescine, cadaverine, histamine, tyramine. This standard applies to alcoholic beverages (wine, beer, wine, etc.), determination of biogenic amines in meat and dairy products, seafood (fish and their products, , shrimp and their products) condiments (vinegar, soy sauce, etc.). This standard limit of quantification: a) alcoholic beverages, vinegar tune ignorant article: serotonin 3��g/kg, ��-phenylethylamine 10��g/kg, putrescine 3��g/kg, cadaverine 7��g/kg, histamine 10��g/kg, buttermilk amine 10��g/kg, spermidine 5��g/kg, spermine 7��g/kg. b) aquatic products, meat, dairy products and beans as raw materials in the spice |
GB 5009.208-2016
NATIONAL STANDARD OF THE
PEOPLE’S REPUBLIC OF CHINA
National Food Safety Standard - Determination of
Biogenic Amine in Food
ISSUED ON: DECEMBER 23, 2016
IMPLEMENTED ON: JUNE 23, 2017
Issued by: National Health and Family Planning Commission of the
People’s Republic of China;
China Food and Drug Administration.
Table of Contents
Foreword ... 3
Method I Liquid Chromatography ... 4
1 Scope ... 4
2 Principle ... 4
3 Reagents and Materials ... 4
4 Instruments and Equipment ... 8
5 Analytical Procedures ... 8
6 Expression of Analytical Result ... 11
7 Precision ... 11
8 Others ... 11
Method II Spectrophotometry ... 13
9 Scope ... 13
10 Principle ... 13
11 Reagents and Materials ... 13
12 Analytical Procedures ... 14
13 Result Calculation ... 15
14 Precision ... 16
15 Others ... 16
Appendix A Liquid Chromatogram of 9 Biogenic Amine Standard Solutions and
Internal Standard Derivatives ... 17
National Food Safety Standard - Determination of
Biogenic Amine in Food
Method I Liquid Chromatography
1 Scope
This Standard specifies the determination method for tryptamine, -phenylethylamine,
putrescine, cadaverine, histamine, octopamine, tyramine, spermidine and spermine
content in food.
This Standard is applicable to the determination of alcohol (wine, beer and yellow rice
wine, etc.), condiments (vinegar and soy sauce), aquatic products (fish and its products;
shrimp and its products), biogenic amine in meat.
2 Principle
Aquatic products (fish and its products; shrimp and its products) and meat: use 5%
trichloroacetic acid to extract sample; use n-hexane to remove fat. After extraction and
purification through trichloromethane-n-butanol (1 + 1) solution, use Dansyl chloride
for derivation. Use C18 chromatographic column for separation. Use high-performance
liquid chromatography-UV detector for detection. Use internal standard method for
quantification.
Alcohol (wine, beer and yellow rice wine, etc.) and sediments (vinegar and soy sauce):
use Dansyl chloride for derivation. Use C18 chromatographic column for separation.
Use high-performance liquid chromatography-UV detector for detection. Use internal
standard method for quantification.
3 Reagents and Materials
Unless it is otherwise stipulated, all reagents used in this Method are analytically pure;
water is Grade-1 water stipulated in GB/T 6682.
3.1 Reagents
3.1.1 Acetonitrile (CH3CN): chromatographically pure.
3.1.2 Acetone (C3H6O): chromatographically pure.
3.1.3 Ether(C4H10O): re-evaporated.
3.2.7 Saturated sodium bicarbonate solution: weigh-take 15 g of sodium bicarbonate;
add 100 mL of water to dissolve it; take the supernatant as the saturated solution.
3.2.8 50 mg/mL sodium glutamate solution: accurately weigh-take 5.0 g of sodium
glutamate; use saturated sodium bicarbonate solution to dissolve it and reach a
constant volume of 100 mL.
3.2.9 0.01 mol/L ammonium acetate solution that contains 1% acetic acid: weigh-take
0.77 g of ammonium acetate; dissolve it in water; then, transfer it into a 1,000 mL
volumetric flask. Add 10 mL of formic acid; use water to reach a constant volume to
the scale.
3.2.10 Mobile phase A: measure-take 100 mL of 0.01 mol/L ammonium acetate
solution that contains 1% acetic acid; add it to 900 mL of acetonitrile.
3.2.11 Mobile phase B: measure-take 900 mL of 0.01 mol/L ammonium acetate
solution that contains 1% acetic acid; add it to 100 mL of acetonitrile.
3.3 Standard Substances
3.3.1 Histamine dihydrochloride (C5H9N32HCl, CAS No.: 56-92-8) standard
substance (purity > 99%).
3.3.2 -phenylethylamine hydrochloride (C8H11NHCl, CAS No.: 64-04-0) standard
substance (purity > 98%).
3.3.3 Tyramine hydrochloride (C8H11NOHCl, CAS No.: 60-19-5) standard substance
(purity > 98%).
3.3.4 Putrescine dihydrochloride (C4H12N22HCl, CAS No.: 333-93-7) standard
substance (purity > 98%).
3.3.5 Cadaverine dihydrochloride (C5H14N22HCl, CAS No.: 1476-39-7) standard
substance (purity > 98%).
3.3.6 Tryptamine hydrochloride (C10H12N2HCl, CAS No.: 61-54-1) standard substance
(purity > 99%).
3.3.7 Spermine tetrahydrochloride (C10H26N44HCl, CAS No.: 306-67-2) standard
substance (purity > 97%).
3.3.8 Spermidine trihydrochloride (C7H19N33HCl, CAS No.: 334-50-9) standard
substance (purity > 97%).
3.3.9 Octopamine hydrochloride (C8H11NO2HCl, CAS No.: 770-05-9) standard
substance (purity > 97%).
3.3.10 1,7-Diaminoheptane (C7H18N2, CAS No.: 646-19-5) internal standard substance
5.1.3.1 Fat removal: transfer-take 10 mL of the above-mentioned sample extract into
a 25 mL test tube with a plug. Add 0.5 g of sodium chloride; conduct vortex oscillation,
till sodium chloride completely dissolves. Then, add 10 mL of n-hexane; conduct vortex
oscillation for 5 min. Place it still for stratification; then, discard the upper layer of
organic phase. Add 10 mL of n-hexane to the lower layer of sample solution to conduct
fat removal one more time.
5.1.3.2 Extraction: transfer-take 5 mL of the above-mentioned post-fat-removal sample
solution into a 10 mL centrifuge tube with a plug. Use 5 mol/L sodium hydroxide
solution (several drops) to adjust pH to around 12.0. Add 5 mL of N-
butanol/trichloromethane (1 + 1) mixing solution. Conduct vortex oscillation for 5 min.
At 5,000 r/min, conduct centrifugation for 5 min. Then, transfer the upper layer of
organic phase into another 10 mL centrifuge tube with a plug. Re-extract the lower
layer of the sample solution; combine the extract. Use N-butanol/trichloromethane (1
+ 1) to dilute to the scale. Take 5 mL of the extract; add 200 L of hydrochloric acid (1
mol/L); thoroughly mix it up. In 40 °C water bath, use nitrogen to blow it to dryness.
Add 1 mL of hydrochloric acid (0.1 mol/L); conduct vortex oscillation. Thoroughly
dissolve the residue. Reserve it for derivation.
5.1.4 Derivation
5.1.4.1 Derivation of sample: successively add 1 mL of saturated sodium bicarbonate
solution, 100 L of sodium hydroxide solution (1 mol/L) and 1 mL of derivatization
reagent to the above-mentioned sample solution, which is reserved for derivation.
Conduct vortex and mixing for 1 min, then, place it in 60 °C constant-temperature water
bath to conduct derivation for 15 min. Then, take it out; add 100 L of sodium glutamate
solution; conduct oscillation mixing. At 60 °C, conduct constant-temperature reaction
for 15 min. Take it out, then, cool it down to room temperature. Add 1 mL of water to
each centrifuge tube; conduct vortex mixing for 1 min. In 40 °C water bath, use nitrogen
to blow it, so as to remove acetone (around 1 mL). Add 0.5 g of sodium chloride;
conduct vortex oscillation, till sodium chloride completely dissolves. Then, add 5 mL of
ether; then, conduct vortex oscillation for 2 min; place it still for stratification. Then,
suck out the upper layer of organic phase (the ether layer); re-extract once. Combine
the ether extract; in 40 °C water bath, use nitrogen to blow it to dryness. Add 1 mL of
acetonitrile to conduct vortex oscillation, so that the residue can completely dissolve.
Use 0.22 m membrane needle filter to filter it into a small sample injection bottle.
Reserve it for determination.
5.1.4.2 Derivation of the standards: respectively transfer-take 1 mL of biogenic amine
standard series solution; place it in a 10 mL test tube with a plug. Successively add
250 L of internal standard working solution (100 mg/L). The following operation is the
same as the steps of sample derivation.
5.2 Alcohol and Condiments (vinegar and soy sauce)
5.2.1 Sample derivation: accurately measure-take 1.0 mL of sample; place it into a 15
Method II Spectrophotometry
9 Scope
This Standard specifies the determination method for histamine content in aquatic
products.
This Standard is applicable to the determination of histamine in aquatic products (fish
and its products; shrimp and its products).
10 Principle
Take trichloroacetic acid as the extraction solution; conduct oscillation extraction.
Through extraction and purification with N-pentanol, histamine and azo reagent trigger
chromogenic reaction. Then, use spectrophotometer for detection; use external
standard method for quantification.
11 Reagents and Materials
Unless it is otherwise stipulated, all reagents used in this Method are analytically pure;
water is Grade-1 water stipulated in GB/T 6682.
11.1 Standards and Reagents
11.1.1 Histamine phosphate (C5H9N32H3PO4).
11.1.2 N-pentanol (C5H12O).
11.1.3 Trichloroacetic acid (C2HCl3O2).
11.1.4 Sodium carbonate (Na2CO3).
11.1.5 Sodium hydroxide (NaOH).
11.1.6 Hydrochloric acid (HCl, 37%).
11.1.7 P-nitroaniline (C6H6N2O2).
11.1.8 Sodium nitrite (NaNO2).
11.2 Preparation of Reagents
11.2.1 Histamine standard stock solution: at 100 °C (± 5 °C), dry histamine phosphate
standard for 2 h. Then, weigh-take 0.2767 g (accurate to 0.001 g) of it, place it in a 50
mL beaker. Use a proper amount of water to completely dissolve it, then, transfer it into
mark them. Store them at 20 °C.
12.2 Sample Analysis
12.2.1 Sample extraction
Accurately weigh-take 10 g (accurate to 0.01 g) of sample, which is already minced
and homogenized. Place it into a 100 mL conical flask with a plug. Add 20 mL of 10%
trichloroacetic acid solution to soak it for 2 h ~ 3 h. Conduct oscillation for 2 min, then,
mix it up. Use filter paper to filter it. Accurately absorb 2.0 mL of the filtrate, then, place
it in a separating funnel. Dropwise add sodium hydroxide solution to adjust pH between
10 ~ 12. Add 3 mL of N-pentanol, then, conduct oscillation extraction for 5 min. Place
it still for stratification. Transfer the N-pentanol extract (the upper layer) to 10 mL scale
test tube. Extract N-pentanol for 3 times; combine the extract; use N-pentanol to dilute
to the scale. Absorb 2.0 mL of N-pentanol extract into separating funnel; add 3 mL of
hydrochloric acid; conduct oscillation extraction. Place it still for stratification. Transfer
the hydrochloric acid extract (the lower layer) to 10 mL scale test tube. Conduct
extraction for 3 times; then, combine the extract; use hydrochloric acid solution to dilute
it to the scale.
12.2.2 Determination
Respectively absorb 0, 0.20, 0.40, 0.60, 0.80 and 1.0 mL of histamine standard working
solution (equivalent to 0, 4.0, 8.0, 12, 16 and 20 g of histamine) and 2.0 mL of sample
extract into a 10 mL colorimetric tube. Add water to 1 mL, then, add 1 mL of
hydrochloric acid solution; mix it up. Add 3 mL of sodium carbonate solution and 3 mL
of azo reagent. Add water to the scale, mix it up; place it for 10 min. Transfer “0” tube
solution into a 1 cm cuvette; adjust the wavelength of the spectrophotometer to 480
nm; adjust the absorbance to “0”. Then, successively test the absorbance of the series
standard solution and the sample solution. Take absorbance A as the vertical axis; take
the mass of histamine as the horizontal axis; draw a standard curve.
13 Result Calculation
The content of histamine in the sample shall be calculated in accordance with Formula
(2):
Where:
X---the content of histamine in the sample, expressed in (mg/100 g);
m1---the mass of histamine which is corresponding with the absorbance value of
histamine in the sample, expressed in (g);
......
GB/T 5009.208-2008
Determination of biogenic amine in foods
ICS 67.040
C53
National Standards of People's Republic of China
Determination of levels of biogenic amines
Posted 2008-11-21
2009-03-01 implementation
People's Republic of China Ministry of Health
Standardization Administration of China released
Foreword
Appendix A of this standard is an informative annex.
This standard is proposed and administered by the People's Republic of China Ministry of Health.
This standard by the People's Republic of China Ministry of Health is responsible for interpretation.
This standard was drafted. Chinese Center for Disease Control Nutrition and Food Safety.
Participated in the drafting of this standard. Beijing Center for Disease Control and Prevention, Yantai University, Henan Drug Control.
The main drafters of this standard. Wu Yongning, Zhao Yunfeng, Li Zhijun, Lu Bin, Li Jingguang, Zhang Lei, Shao Bing, Miao Hong.
Determination of levels of biogenic amines
1 Scope
This standard specifies the foods tryptamine, β- phenylethylamine, putrescine, cadaverine, histamine, tyramine, spermidine and spermine content determination.
This standard applies to alcohol (wine, beer, wine, etc.), spices (vinegar, soy sauce, etc.), seafood (fish and their products, and shrimp
Products), meat and dairy products Determination of biogenic amines.
The standard quantitative limit.
a) wine, vinegar condiment. tryptamine 3μg/kg, β- phenylethylamine 10μg/kg, putrescine 3μg/kg, cadaverine 7μg/kg, histamine
10μg/kg, tyramine 10μg/kg, spermidine 5μg/kg, spermine 7μg/kg.
b) aquatic products, meat, dairy products and beans as raw material to seasoning. tryptamine 15μg/kg, β- phenylethylamine 40μg/kg, putrescine
15μg/kg, cadaverine 35μg/kg, histamine 50μg/kg, tyramine 25μg/kg, spermidine 25μg/kg, spermine 35μg/kg.
Principle 2
With 1,7-amino-heptane as an internal standard, with 5% trichloroacetic acid solution to extract, shaking extraction with hexane to remove fat, through trichlorosilane
Alkyl - n-butanol (1 + 1) liquid-liquid extraction purified to dansyl chloride derived agents, 60 ℃ derivative 30min, HPLC C18 column
Separation, UV detector, internal standard.
3 Reagents and materials
Unless otherwise specified in this standard reagents used were of analytical grade.
3.1 methanol (CH4O). chromatography.
3.2 acetone (C3H6O). chromatography.
3.3 ether (C4H10O). redistilled.
3.4 n-butanol (C4H10O).
3.5 chloroform (CHCl3).
3.6 n-hexane (C6H14). chromatography.
3.7 sodium glutamate (C5H8NNaO4).
3.8 sodium bicarbonate (NaHCO3).
3.9 Sodium chloride (NaCl).
3.10 Sodium hydroxide (NaOH).
3.11 hydrochloric acid (HCl).
3.12 TCA (C2HCl3O2).
3.13 histamine hydrochloride (histaminedihydrochloride, C5H9N3 · 2HCl) standards (purity > 99%, when calculating conversion off
Hydrochloride, C5H9N3/C5H9N3 · 2HCl = 155/184).
3.14 β- phenylethylamine (β-phenylethylamine, C8H11N) standards (purity > 98%).
3.15 tyramine hydrochloride (tyraminehydrochloride, C8H12ClNO) standards (purity > 99%, when calculating conversion off hydrochloride,
C8H11NO/C8H11NO · HCl = 137/174).
3.16 putrescine (putrescine, C4H12N2) standards (purity > 98%).
3.17 cadaverine (cadaverine, C5H14N2) standards (purity > 98%).
3.18 tryptamine (tryptaminehydrochloride, C10H12N2) standards (purity > 99%).
3.19 spermidine (spermidine, C7H19N3) standards (purity > 97%).
......
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