GB/T 22224-2008 PDF English
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Determination of dietary fiber in foods -- Enzymatic gravimetric method and enzymatic gravimetric method -- Liquid chromatography
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GB/T 22224-2008: Determination of dietary fiber in foods -- Enzymatic gravimetric method and enzymatic gravimetric method -- Liquid chromatography
---This is an excerpt. Full copy of true-PDF in English version (including equations, symbols, images, flow-chart, tables, and figures etc.), auto-downloaded/delivered in 9 seconds, can be purchased online: https://www.ChineseStandard.net/PDF.aspx/GBT22224-2008
NATIONAL STANDARD OF THE
PEOPLE’S REPUBLIC OF CHINA
ICS 67.050
X 04
Determination of dietary fiber in foods - Enzymatic
gravimetric method and enzymatic gravimetric
method-liquid chromatography
Issued on. MAY 16, 2008
Implemented on. OCTOBER 01, 2008
Issued by. General Administration of Quality Supervision, Inspection and
Quarantine;
Standardization Administration of the People's Republic of
China.
Table of Contents
Foreword... 4
1 Method One - Enzymatic-gravimetric method... 5
1.1 Scope... 5
1.2 Normative references... 5
1.3 Terms and definitions... 5
1.4 Method summary... 6
1.5 Reagents and solutions... 6
1.6 Apparatus and equipment... 8
1.7 Sample preparation... 9
1.8 Analysis steps... 9
1.9 Result calculation... 11
1.10 Tolerance... 12
2 Method Two - Enzymatic-gravimetric method and liquid chromatography
method... 12
2.1 Scope... 12
2.2 Normative references... 13
2.3 Terms and definitions... 13
2.4 Method summary... 13
2.5 Reagents and solutions... 14
2.6 Apparatus and equipment... 14
2.7 Sample preparation... 15
2.8 Analysis steps... 15
2.9 Result calculation... 18
2.10 Tolerance... 20
1 Method One - Enzymatic-gravimetric method
1.1 Scope
Method One of this Standard specifies the conditions and detailed analysis
steps for the determination of total, soluble and insoluble dietary fiber in foods
by enzymatic-gravimetric method.
Method One of this Standard is applicable to the determination of total, soluble
and insoluble dietary fiber in cereals, vegetables and fruits as well as their
products. But it is not applicable to the determination of dietary fiber in foods
containing low-molecular-mass resistant maltodextrin, oligofructose, galacto-
oligosaccharides, polyglucose and resistant starch.
1.2 Normative references
The provisions in following documents become the provisions of this Standard
through reference in this Standard. For dated references, the subsequent
amendments (excluding corrigendum) or revisions do not apply to this Standard,
however, parties who reach an agreement based on this Standard are
encouraged to study if the latest versions of these documents are applicable.
1.3 Terms and definitions
For the purposes of Method One of this Standard, the following terms and
definitions apply.
1.3.1 dietary fiber
the sum of carbohydrates and their analogues that have anti-digestive
properties, which cannot be absorbed by the human small intestine but can be
partially or completely fermented in colon
1.3.2 total dietary fiber; TDF
including insoluble dietary fiber (IDF) and high-molecular-mass soluble dietary
fiber (SDF) that can be precipitated in ethanol
1.4 Method summary
Use thermostable α-amylase, protease and amyloglucosidase to perform
enzymatic digestion of the dried sample. Use ethanol to precipitate enzymatic
hydrolysate. Filter.
1.5 Reagents and solutions
Unless otherwise indicated, in analysis, it shall only use the confirmed
analytically-pure reagent AND distilled water or deionized water or equivalent-
pure water.
1.5.1 95% ethanol.
1.5.2 Thermostable α-amylase solution. CAS 9000-85-5, IUS 3.2.1.1; cannot
contain glycerol as stabilizer. Store at 0°C ~ 5°C in refrigerator.
1.5.2.1 Enzyme activity representation 1.Starch is substrate. It is represented
by Nelson / Somogyi reducing sugar -- 10,000 units/mL + 1,000 units/mL (1
enzyme activity unit is defined as. at 40°C, when pH6.5, the amount of enzyme
required to release 1 μmol of reducing sugar per minute).
1.5.5 Pickled diatomite. CAS 68855-54-9.Take 200 g of diatomite in 600 mL of
hydrochloric acid (HCl. H2O = 1.4, volume ratio). Soak overnight. Filter. Use
distilled water to wash until the filtrate is neutral. Place in 525°C ± 5°C muffle
furnace to burn the ash. Leave it for use.
1.5.6 2-(N-morpholino)-sulfonate ethane (MES). CAS 4432-31-9, purity >99.5%.
1.5.7 Trihydroxymethyl aminomethane (TRIS). CAS 77-86-1, purity >99%.
1.5.8 MES-TRIS buffer (0.05 mol/L). Weigh 19.52 g of MES and 12.2 g of TRIS.
Use 1.7 L of distilled water to dissolve. Use 6 mol/L sodium hydroxide to adjust
pH to 8.2 ± 0.1.Add water to dilute to 2 L (attention. pH is 8.2 at 24°C; pH is 8.3
at 20°C; pH is 8.1 at 28°C. Be sure to adjust pH according to the temperature.
The deviation between 20°C and 28°C is corrected by interpolation method).
1.5.9 Hydrochloric acid solution (0.561 mol/L). Take 93.5 mL of 6 mol/L
hydrochloric acid. Add 700 mL of water. Mix well. Use water to set volume to 1
L.
1.5.10 Petroleum ether. Boiling range is 30°C ~ 60°C.
1.5.11 Acetone.
1.5.12 Sodium hydroxide.
1.6 Apparatus and equipment
1.6.1 High-type beaker without diversion inlet. 400 mL or 600 mL.
1.6.2 Crucible. with rough sintered glass plate; the aperture is 40 µm ~ 60 µm.
Ash the cleaned crucible in muffle furnace at 525°C for 6 h. Take it out when
the furnace temperature reduces below 130°C. Soak in potassium dichromate
lotion for 2 h. Respectively use water and distilled water to rinse clean. In the
end, use 15 mL of acetone to rinse then air-dry it.
1.6.3 Vacuum solvent filter. vacuum pump or aspirator with adjustment device.
1L suction-filtration bottle of which there is a suction-filtration mouth on the side
wall; there is a rubber stopper matching with the suction-filtration bottle. It is
used for the suction-filtration of enzymatic hydrolysate.
1.6.4 Constant-temperature oscillating water bath. 95°C ~ 100°C.
1.6.5 Analytical balance. resolution is 0.1 mg.
1.6.6 Balance (platform scale). measuring range is 4,000 g; resolution is 0.1 g.
1.6.7 Muffle furnace. 525°C ± 5°C.
1.6.8 Oven. 105°C, 130°C ± 3°C.
1.6.9 Vacuum drying oven.
1.6.10 Dryer. silica or equivalent desiccant.
1.6.11 pH meter. with temperature compensation function; accuracy is ±0.1.
1.6.12 Micro-Kjeldahl.
1.6.13 Pipette. 100 µL, 5 mL; disposable pipette tip.
1.7 Sample preparation
1.7.1 Food with fat content less than 10%
Take well-mixed sample at 70°C vacuum to dry overnight. Place in the dryer to
cool. After the dry sample is smashed, sieve through 0.3mm ~ 0.5mm sieve. If
the sample cannot be heated, then lyophilize it first. Smash and sieve. Store
the smashed and sieved dry sample in the dryer for use.
1.7.3 Food with high sugar content
Take an appropriate amount of sample. Add 10 mL of 85% ethanol per gram of
sample. Process the sample 2 ~ 3 times to de-sugar. Dry overnight at 40°C.
Store the smashed-sieved dry sample in the dryer for use.
1.8 Analysis steps
1.8.1 Determination of moisture content
Determine the moisture content in sample according to GB/T 5009.3-2003.Use
it for result calculation.
1.8.2 Enzymatic hydrolysis
1.8.2.1 Accurately weigh double samples (ms1 and ms2), 1 g for each. The mass
difference between two samples is ≤0.005 g, to the nearest of 0.1 mg. Place in
400mL or 600mL high-type beaker (1.6.1). Prepare double blank samples at the
same time. Add 40 mL of pH8.2 MES-TRIS buffer into each beaker.
1.8.3 Determination
1.8.3.1 Determination of total dietary fiber
1.8.3.1.1 Precipitation. In each sample, add 225 mL (the volume after
preheating) of 95% ethanol that has been preheated to 60°C. The volume ratio
of ethanol to sample solution is 4.1.Take the beaker out. Cover aluminum foil.
Precipitate at room temperature for 1 h. It is recommended to modify as. Weigh
the mass of enzymatic hydrolysate. Use balance (1.6.6) to add 4 times mass
95% ethanol that has been preheated to 60°C. Precipitate overnight in 4°C
refrigerator.
1.8.3.2 Determination of insoluble dietary fiber
1.8.3.2.1 Weigh the sample mass according to 1.8.2.1.Perform enzymatic
hydrolysis according to 1.8.2.2 ~ 1.8.2.6.
1.8.3.2.2 Filtration and washing. Transfer all sample enzymatic hydrolysate to
crucible to filter.
1.8.3.2.3 Determine protein and ash according to 1.8.3.1.4.
1.8.3.3 Determination of soluble dietary fiber
2 Method Two - Enzymatic-gravimetric method and
liquid chromatography method
2.1 Scope
Method Two of this Standard specifies the conditions and detailed analysis
steps for determination of total dietary fiber in foods containing low-molecular-
mass resistant maltodextrin by enzymatic-gravimetric method and liquid
chromatography method.
2.2 Normative references
The provisions in following documents become the provisions of this Standard
through reference in this Standard. For dated references, the subsequent
amendments (excluding corrigendum) or revisions do not apply to this Standard,
however, parties who reach an agreement based on this Standard are
encouraged to study if the latest versions of these documents are applicable.
2.3 Terms and definitions
For the purposes of Method Two of this Standard, the following terms and
definitions apply.
2.4 Method summary
Take two samples. Under the sequential actions of thermostable α-amylase,
protease and amyloglucosidase, perform enzymatic hydrolysis to make starch,
protein in sample into dissolved small molecules.
2.5 Reagents and solutions
Unless otherwise indicated, in analysis, it shall only use the confirmed
analytically-pure reagent AND distilled water or deionized water or equivalent-
pure water.
2.5.1 ~ 2.5.12 Same with 1.5.1 ~ 1.5.12 of Method One.
2.6 Apparatus and equipment
Laboratory routine apparatus and the followings.
2.6.1 ~ 2.6.13 Same with 1.6.1 ~ 1.6.13 of Method One.
2.6.17 Gel protection column. 6.0 mm × 40 mm, 6 μm.
2.6.18 Gel chromatographic column. 7.8 mm × 300 mm, 6 μm; two pieces.
2.7 Sample preparation
2.7.1 Same with 1.7.1 of Method One.
2.7.2 Same with 1.7.2 of Method One. But the foods that contain high sugar
content may not be de-sugared.
2.8 Analysis steps
2.8.1 Determination of moisture content
Same with 1.8.1 of Method One.
2.8.2 Enzymatic hydrolysis
2.8.2.4 Same with 1.8.2.4 of Method One.
2.8.2.5 Same with 1.8.2.5 of Method One.
2.8.2.6 Same with 1.8.2.6 of Method One. The amount of amyloglucosidase
used is 300 μL.
2.8.3 Enzymatic gravimetric method
It is used to determine the total content of insoluble dietary fiber (IDF) and high-
molecular-mass soluble dietary fiber (SDF).
2.8.3.1 Precipitation. same with 1.8.3.1.1.
2.8.3.2 Filtration. same with 1.8.3.1.2.
2.8.3.3 Washing. Use 20 mL of 78% ethanol to wash high-type beaker 3 times.
Under vacuum conditions, wash off the residues in crucible.
2.8.3.6 Determination of ash. Take one residue from double samples.
Determine the ash in the sample according to GB/T 5009.4-2003.
2.8.3.7 Determination of protein. Take the other residue from double samples.
Determine the protein content in the sample according to GB/T 5009.5-2003.
2.8.4 Liquid chromatography
Use high performance liquid chromatography to determine the content of low-
molecular-mass of resistant maltodextrin in sample.
......
Source: Above contents are excerpted from the full-copy PDF -- translated/reviewed by: www.ChineseStandard.net / Wayne Zheng et al.
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