CN106970041A - A kind of near-infrared assay method of the big aggressiveness content of wheat flour Insoluble glutenin - Google Patents
A kind of near-infrared assay method of the big aggressiveness content of wheat flour Insoluble glutenin Download PDFInfo
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- 108010050792 glutenin Proteins 0.000 title claims abstract description 51
- 235000013312 flour Nutrition 0.000 title claims abstract description 30
- 241000209140 Triticum Species 0.000 title claims abstract description 22
- 235000021307 Triticum Nutrition 0.000 title claims abstract description 22
- 238000003556 assay Methods 0.000 title claims abstract description 22
- 238000012360 testing method Methods 0.000 claims abstract description 21
- 239000000126 substance Substances 0.000 claims abstract description 19
- 239000007788 liquid Substances 0.000 claims abstract description 15
- 238000000034 method Methods 0.000 claims description 25
- 239000012530 fluid Substances 0.000 claims description 14
- 239000008363 phosphate buffer Substances 0.000 claims description 13
- 230000000694 effects Effects 0.000 claims description 11
- 238000004458 analytical method Methods 0.000 claims description 10
- 238000004611 spectroscopical analysis Methods 0.000 claims description 10
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims description 9
- 238000002329 infrared spectrum Methods 0.000 claims description 9
- 238000001228 spectrum Methods 0.000 claims description 9
- 238000004587 chromatography analysis Methods 0.000 claims description 8
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 claims description 6
- 238000005259 measurement Methods 0.000 claims description 5
- 239000006228 supernatant Substances 0.000 claims description 5
- 239000010453 quartz Substances 0.000 claims description 4
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N silicon dioxide Inorganic materials O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims description 4
- 239000004677 Nylon Substances 0.000 claims description 3
- 238000005119 centrifugation Methods 0.000 claims description 3
- 238000002512 chemotherapy Methods 0.000 claims description 3
- 239000012528 membrane Substances 0.000 claims description 3
- 229920001778 nylon Polymers 0.000 claims description 3
- 238000001556 precipitation Methods 0.000 claims description 3
- 230000009514 concussion Effects 0.000 claims description 2
- 239000000047 product Substances 0.000 claims description 2
- 238000001514 detection method Methods 0.000 abstract description 5
- 239000000203 mixture Substances 0.000 abstract description 2
- 238000012795 verification Methods 0.000 abstract 1
- 239000000523 sample Substances 0.000 description 34
- 239000000243 solution Substances 0.000 description 12
- 235000013339 cereals Nutrition 0.000 description 9
- 108090000623 proteins and genes Proteins 0.000 description 8
- 235000018102 proteins Nutrition 0.000 description 7
- 102000004169 proteins and genes Human genes 0.000 description 7
- 230000003595 spectral effect Effects 0.000 description 7
- NBIIXXVUZAFLBC-UHFFFAOYSA-N phosphoric acid Substances OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 6
- 108010068370 Glutens Proteins 0.000 description 5
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 4
- 239000000463 material Substances 0.000 description 4
- 239000011159 matrix material Substances 0.000 description 4
- 235000012239 silicon dioxide Nutrition 0.000 description 4
- 238000002835 absorbance Methods 0.000 description 3
- OHJMTUPIZMNBFR-UHFFFAOYSA-N biuret Chemical compound NC(=O)NC(N)=O OHJMTUPIZMNBFR-UHFFFAOYSA-N 0.000 description 3
- 239000000872 buffer Substances 0.000 description 3
- 238000004737 colorimetric analysis Methods 0.000 description 3
- 235000021312 gluten Nutrition 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 244000098338 Triticum aestivum Species 0.000 description 2
- 238000009395 breeding Methods 0.000 description 2
- 230000001488 breeding effect Effects 0.000 description 2
- 238000003391 densitometric scan Methods 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 238000002270 exclusion chromatography Methods 0.000 description 2
- 238000001502 gel electrophoresis Methods 0.000 description 2
- 229910052739 hydrogen Inorganic materials 0.000 description 2
- 238000005192 partition Methods 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- BDERNNFJNOPAEC-UHFFFAOYSA-N propan-1-ol Chemical compound CCCO BDERNNFJNOPAEC-UHFFFAOYSA-N 0.000 description 2
- 235000019624 protein content Nutrition 0.000 description 2
- 239000002699 waste material Substances 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- HNSDLXPSAYFUHK-UHFFFAOYSA-N 1,4-bis(2-ethylhexyl) sulfosuccinate Chemical compound CCCCC(CC)COC(=O)CC(S(O)(=O)=O)C(=O)OCC(CC)CCCC HNSDLXPSAYFUHK-UHFFFAOYSA-N 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 238000004497 NIR spectroscopy Methods 0.000 description 1
- RCEAADKTGXTDOA-UHFFFAOYSA-N OS(O)(=O)=O.CCCCCCCCCCCC[Na] Chemical compound OS(O)(=O)=O.CCCCCCCCCCCC[Na] RCEAADKTGXTDOA-UHFFFAOYSA-N 0.000 description 1
- 238000000862 absorption spectrum Methods 0.000 description 1
- 235000009120 camo Nutrition 0.000 description 1
- 244000213578 camo Species 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 238000002790 cross-validation Methods 0.000 description 1
- 238000000354 decomposition reaction Methods 0.000 description 1
- 230000001934 delay Effects 0.000 description 1
- 235000013399 edible fruits Nutrition 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 238000011010 flushing procedure Methods 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 238000005227 gel permeation chromatography Methods 0.000 description 1
- 230000005283 ground state Effects 0.000 description 1
- 238000003306 harvesting Methods 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 208000000509 infertility Diseases 0.000 description 1
- 230000036512 infertility Effects 0.000 description 1
- 231100000535 infertility Toxicity 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 238000012417 linear regression Methods 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 238000011068 loading method Methods 0.000 description 1
- 238000000691 measurement method Methods 0.000 description 1
- 238000001320 near-infrared absorption spectroscopy Methods 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 230000031787 nutrient reservoir activity Effects 0.000 description 1
- 238000006116 polymerization reaction Methods 0.000 description 1
- 238000004445 quantitative analysis Methods 0.000 description 1
- 238000003044 randomized block design Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000001542 size-exclusion chromatography Methods 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- -1 sodium alkyl sulfate Chemical class 0.000 description 1
- 230000007704 transition Effects 0.000 description 1
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
- G01N21/31—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry
- G01N21/35—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry using infrared light
- G01N21/359—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry using infrared light using near infrared light
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/34—Purifying; Cleaning
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- G—PHYSICS
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/04—Preparation or injection of sample to be analysed
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Abstract
The invention belongs to agricultural product composition detection technical field, and in particular to a kind of near-infrared assay method of big aggressiveness content of wheat flour Insoluble glutenin and its application.It is a feature of the present invention that：After the buffered liquid of wheat flour is extracted, the liquid testing sample containing the big aggressiveness of Insoluble glutenin is obtained.Testing sample is while true chemical value is determined, and reflected formula near infrared spectrometer scanning gathers primary light spectrogram.The near-infrared for setting up the big aggressiveness content of flour Insoluble glutenin by stoichiometry software and PLS predicts calibrating patterns.According to model parameter and application verification, the forecast model has outstanding feature accurately and fast with low cost.
Description
Technical field
The application is related to agricultural product composition detection technical field, particularly a kind of big aggressiveness of wheat flour Insoluble glutenin
The near-infrared assay method of content.
Background technology
The anharmonicity of near infrared spectrum (Near-infrared spectroscopy, NIRS) based on molecular vibration, leads
Cause molecular vibration from ground state to being produced during high energy order transition, wave-length coverage can record hydric group generally in 800~2500nm
The frequency multiplication and sum of fundamental frequencies of the vibrations such as C-H, O-H, N-H, S-H, P-H absorb, the association set up between absorption spectrum and content of material.It is non-
Often suitable for the content analysis of the carbon-hydrogen organics such as the moisture content analysis and albumen, carbohydrate, lipid of checking matter.
Glutelin in wheat seed and flour generally exists in polymeric form, and wherein the degree of polymerization is higher, 12
The part for being difficult to be extracted in sodium alkyl sulfate phosphate buffer is referred to as the big aggressiveness of Insoluble glutenin
(Unextractablepolymeric protein, UPP).UPP only accounts for about the 50% of wheat gluten total amount, and storage protein is total
About the 5% of amount, but be in extremely significant positive correlation with gluten strength, UPP contents are mainly influenceed by genotype, and it is to gluten quality
There is conclusive effect (ZhangP P, He Z H, XiaX C, et al.Effect ofpercent SDS- with food quality
Unextractable polymeric protein (%UPP) on end-use quality in Chinese
breadwheat cultivars.Cereal Chemistry,2008,85:696-700；Zhu J,KhanK.Effects
ofgenothpe and environment on gluteninpolymers andbreadmaking quality.Cereal
Chem,2001,78:125-130；
GuptaRB,Khan K,MacRitchie F.Biochemical basis offlourproperties
inbread wheats.I.Effects ofvariation in the quantity and size distribution
ofpolymeric protein.J Cereal Sci,1993,18:23-41).Therefore, how fast and accurately quantitative analysis
UPP turns into the important content that wheat breed breeding generation is selected.
Conventional Glutenin macropolymer analysis method includes at present：Biuret colorimetric method (Liu Li, Zhouyang, He Zhonghu, etc.
.Glu-1 the influence Acta Agronomica Sinicas with Glu-3 allelic variations to Insoluble glutenin content, 2004,30:1086-1092), third
Alcohol partition method (Bean S R, Lyne RK, TilleyKA, et al.A rapidmethod for quantitation
ofinsoluble polymeric proteins in flour.CerealChem,1998,75:374-379；Fu B X,
SapirsteinH D.Procedure for isolating monomeric proetins andpolymeric
glutenin ofwheat flour.Cereal Chem,1996,73:143-152), multilayer gel electrophoresis densitometric scan method
(Zhu J,Khan K.Effects ofgenothpe and environment on gluteninpolymers
andbreadmaking quality.Cereal Chem,2001,78:125-130) with exclusion chromatography (Zhang P P, He
Z H, Xia X C, et al.Effect ofpercent SDS-unextractable polymeric protein (%UPP)
on end-use quality in Chinese bread wheat cultivars.Cereal Chemistry,2008,85:
696-700；GuptaR B,Khan K,MacRitchie F.Biochemical basis offlour properties
inbread wheats.I.Effects of variation in the quantity and size distribution
ofpolymeric protein.J Cereal Sci,1993,18:23-41)。
Though biuret colorimetric method and propyl alcohol partition method method be simple, the not Glutenin macropolymer of its practical measurement contains
Amount, but the polymeric total amount of glutelin, and also be present the poor repeatability between parallel samples in biuret colorimetric method, waste time and energy
The problem of.Multilayer gel electrophoresis densitometric scan method continuous mode and technical sophistication, the dyeing-decolorzing requirement to gel are strict, into
This is higher, wastes time and energy.Exclusion chromatography is current most accurate method, but this method equipment and consumptive material are expensive, it is necessary to special
Technical staff, be used for the higher quantitative study of requirement, but the application in breeding assisted Selection is less.At present, using near
The research that infrared spectrum technology is measured to Glutenin macropolymer content has no report.
The content of the invention
In view of the above-mentioned problems, the present invention, which is established, can determine the near infrared spectrum of wheat flour Glutenin macropolymer content
Assay method (model), this method sample preparation is simple, with low cost, and measurement accuracy is high.What the present invention was realized in：
A kind of near-infrared assay method of the big aggressiveness content of wheat flour Insoluble glutenin, it is comprised the following steps that：1)
Prepare Glutenin macropolymer sample：The big aggressiveness fluid sample of albumen is prepared using SDS- phosphate buffer extract solutions, it is standby；
Specific extracting method is to weigh 20mg flour samples respectively in 2.0mL centrifuge tubes, adds 1.6mL SDS- phosphoric acid and delays
Fliud flushing extract solution shakes 10min, 17000g centrifugation 5min, abandons supernatant；Add 1.6mL SDS- phosphoric acid buffers again in precipitation
Liquid extract solution, 20s is extracted using Ultrasonic cell smash, and then 17000g centrifuges 5min, the supernatant obtained as paddy
The big aggressiveness fluid sample of albumen；
2) step 1 is taken using 0.45 μm of nylon membrane (water system) syringe filters respectively) the Glutenin macropolymer liquid that obtains
Sample 200uL carries out chromatography, obtains chemical assay result；
3) take step 1 respectively) obtain Glutenin macropolymer 1.0~1.2mL of fluid test sample, gather spectroscopic data；
Spectral scan interval 950-1650nm, scanning times 10 times, temperature room temperature, resolution ratio 5nm；
4) deflected secondary air is used, to step 2) obtain chemical assay result and step 3) obtain spectroscopic data
Analysis is calculated, forecast model is set up；The coefficient of determination of the model is R2=0.89, the calibration standard deviation of model is RMSECV=
36.40；
5) Unscrambler chemo metric softwares are used, the calibrating die of prediction is set up with reference to PLS method
Type；
Spectrum baseline in measurement process is eliminated using the smooth Pretreated spectra modes of first derivative+S-G to drift about to setting up
The influence that the calibrating patterns of prediction are caused, with the standard deviation of validation-cross and the coefficient of determination R of institute's established model2For evaluation and foreca
Calibrating patterns effect major parameter；
6) using and step 1) identical method prepares the Glutenin macropolymer fluid sample of testing sample, utilizes step 5)
Calibrating patterns of the prediction of acquisition, and according to step 3) methods described carries out near-infrared spectrum analysis to testing sample, that is, obtains
The content of the big aggressiveness of testing sample Insoluble glutenin.
Further, in the near-infrared assay method of the big aggressiveness content of wheat flour Insoluble glutenin of the present invention, step
Rapid 1) described SDS- phosphate buffers extract solution refers to, the 0.5M phosphate buffers of the 0.5%SDS containing mass volume ratio, pH value
6.90。
Further, in the near-infrared assay method of the big aggressiveness content of wheat flour Insoluble glutenin of the present invention, step
The extracting parameter of rapid 1) described Ultrasonic cell smash isProbe, output work 10W.
Further, in the near-infrared assay method of the big aggressiveness content of wheat flour Insoluble glutenin of the present invention, step
Rapid 2) described chromatography refers to that, using high performance liquid chromatograph, Biosep S4000 gel chromatographic columnses, mobile phase is containing volume
Than 50% acetonitrile solution (v/v) of 0.05% trifluoroacetic acid, flow velocity 0.5mL/min.
Further, in the near-infrared assay method of the big aggressiveness content of wheat flour Insoluble glutenin of the present invention, step
During rapid 3) the collection spectroscopic data, sensing equipment is Perten DA7200 diode array near infrared spectrometers, fills sample hardware
For liquid quartz sample cell, thickness is 1mm.
Compared to existing measuring method, the forecast model provided using the application is to the big aggressiveness of flour Insoluble glutenin
Determine, with outstanding feature accurately and fast with low cost, it is easy to popularization and application.
Brief description of the drawings
Fig. 1 is that the big aggressiveness near-infrared of Insoluble glutenin predicts in calibrating patterns that chemical score is related to spectral predicted value
Property analysis chart.
Fig. 2 is Glutenin macropolymer fluid test sample (extract solution) atlas of near infrared spectra.
Embodiment
Technical scheme is described in detail in following examples, is illustrative purpose, is not to this hair
The limitation of bright protection domain.
Material source/collocation method/the equipment and running parameter being related in embodiment：
1st, the flour samples sources that near-infrared forecast model and model are used：Applicant voluntarily collects Jiangsu, Anhui, mountain
120 parts of the wheat breed (being) in the provinces such as east, Henan, Shaanxi, Sichuan, is planted in 2014-2015 years in agriculture section of Jiangsu Province
Institute proving ground.
Field test uses RANDOMIZED BLOCK DESIGN, 3 row areas, the long 1.5m of row, 2 repetitions, and time of infertility amount of nitrogen is about
225kg.hm-1, it is ripe after harvest seed samples.Two are repeated after seed samples mixed in equal amounts, Brabender is used
Quadrumat Junior Laboratory are according to standardization program powder, flour extraction about 60%.
These sample material protein contents and the big aggressiveness content variation of Insoluble glutenin are abundant (being shown in Table 1), Xia Shushi
Apply in example, randomly selecting in these samples 100 parts is used to model (embodiment 1), and 20 parts are used for model checking (embodiment 2).
The gel chromatography chemical score and spectral predicted value Basic Statistics and Tables of the modeling sample collection of table 1
| Parameter | Average value | Maximum | Minimum value |
| Protein content (%, butt) | 11.25 | 15.03 | 8.54 |
| The big aggressiveness content chemical score (AU/mg) of Insoluble glutenin | 1108.37 | 1805.57 | 854.55 |
2nd, following embodiment near infrared spectra collections or measure main equipment are Perten DA7200 diode array near-infrareds
Spectrometer (Perten Instr Inc. of Sweden), it is liquid quartz sample cell to determine fashionable dress sample hardware, and thickness is 1mm.
3rd, it is the high performance liquid chromatographs of Dionex UltiMate 3000, chromatogram that following embodiment chemical scores, which determine main equipment,
Post is Biosep S4000 gel chromatographic columnses.Call parameter during measure is：Mobile phase is containing 0.05% (v/v) trifluoroacetic acid
50% acetonitrile solution (v/v), flow velocity 0.5mL/min.Chemical score unit conversion is AU/mg, represents paddy egg in every milligram of flour
The absorbance (AU) of white big aggressiveness.
4th, SDS- phosphate buffers extract solution：0.5M phosphoric acid buffers containing 0.5%SDS (w/v, lauryl sodium sulfate)
Liquid, pH 6.90.
The foundation of the big aggressiveness assay near-infrared model of the Insoluble glutenin of embodiment 1
1st, Glutenin macropolymer testing sample is prepared：Randomly select 100 parts of flour samples, every part accurate weigh 20mg in
2.0mL centrifuge tubes, add 1.6mL SDS- phosphate buffer extract solutions room temperature concussion 10min, 17000g centrifugation 5min, abandon
Clearly；Again in precipitation add 1.6mL SDS- phosphate buffer extract solutions, using Ultrasonic cell smash (Visit
Head, 10W) 20s is extracted, then 17000g centrifuges 5min, and the supernatant obtained is Glutenin macropolymer fluid test sample.
2nd, chromatography determines chemical score：200uL steps 1 are taken to obtain using 0.45 μm of nylon membrane (water system) syringe filters
Fluid test sample carry out chromatography, chemical score range of variation is wider, is 854.55AU/mg~1805.57AU/mg.(tool
Body measurement method is referring to GuptaRB, Khan K, MacRitchie F.Biochemical basis offlourproperties
inbreadwheats.I.Effects ofvariation in the quantity and size distribution
ofpolymeric protein.J Cereal Sci,1993,18:23-41；Larroque O R,Bekes F.Rapid
size-exclusion chromatography analysis ofmolecular size distribution forwheat
endospermprotein.Cereal Chem,2000,77:451-453.), the chemical assay knot of 100 parts of samples of the present embodiment
Fruit is shown in Table 2.
3rd, modeling and spectral collection：Chemical score is carried out in step 2 to determine simultaneously, and Glutenin macropolymer liquid is directly taken respectively
1.0~1.2mL of testing sample, the spectroscopic data of sample is gathered using PertenDA7200 liquid quartzs sample cell.Spectral scan
Interval 950-1650nm, scanning times 10 times, temperature room temperature, resolution ratio 5nm.100 parts of Glutenin macropolymer liquid of the present embodiment
The near infrared spectrum data that testing sample (extract solution) is obtained is as shown in Figure 2.
4th, the foundation of the big aggressiveness near-infrared rating model of Insoluble glutenin：By to step 2 chemical score (table 2) and step
The calculating analysis modeling of 3 spectroscopic datas.
Modeling software used in the present embodiment is The UnscramblerX (v10.3, CAMO), and spectroscopic data is shown in Fig. 2,
The modeling method used is offset minimum binary (PLS) method, and modeling wave band is 950-1650nm.
PLS carries out principal component decomposition to spectral information X and concentration information Y matrixes simultaneously in computing, and uses main gene
Returned, model contains the concentration information of spectral information and sample index of correlation；PLS's comprises the following steps that：
First, X and Y matrixes are decomposed, its model is：
Y=UQT+EY (1)
X=TPT+EX (2)
In formula (1), (2), T and P are respectively the score and loading matrix of X matrix, and U and Q are respectively Y score load moment
Battle array, EXAnd EYRemaining residual matrix when respectively with PLS models fitting X and Y；
Then, T and U are made into linear regression：
U=TB (3)
B=(TTT)-1TTY (4)
In prediction, unknown sample matrix X is obtained according to load battle array P firstIt is unknownScore battle array TIt is unknown, then
Concentration prediction value is obtained by formula (5)：
YIt is unknown=TIt is unknownBQ (5)
5th, corresponding forecast model is obtained, using the forecast model to spectroscopic data backward prediction, predicted value (table is obtained
2)。
The chromatography chemical score and model predication value (AU/mg) of 2 100 parts of modeling flour samples of table
The coefficient of determination of the model is R2=0.89, and the validation-cross standard deviation of model is RMSECV=36.40, is only
The 3.28% of modeling sample average.
As can be seen from Table 2, the model accurately detects the big aggressiveness of Insoluble glutenin in SDS phosphoric acid buffer extract solutions
Content.The major absorbance peak of testing sample is located in the range of 1400~1600nm (as shown in Figure 2), a mainly N-H frequency multiplication,
C-H combination frequency and an O-H frequency multiplication absorb.Meanwhile, from figure 2 it can be seen that each component near infrared spectrum region
Absworption peak overlaps each other, therefore uses Unscrambler chemo metric softwares, sets up and predicts with reference to PLS method
Calibrating patterns.
Before modeling, measurement process is eliminated using first derivative+S-G smooth (smooth 7 points of points) Pretreated spectra mode
The influence that middle spectrum baseline drift is caused to modeling.With standard deviation (the Standarderror ofcross of validation-cross
Validation, SECV) and institute's established model the coefficient of determination (R2) it is the major parameter for evaluating modeling effect.
The application of the big aggressiveness assay near-infrared model of the Insoluble glutenin of embodiment 2
In addition choose have neither part nor lot in embodiment 1 modeling 20, sample, using with the step 1 identical sample preparation of embodiment 1
Method obtains Glutenin macropolymer fluid test sample.
Using identical DA7200 near-infrared analyzers in embodiment 1 and liquid quartz sample cell, the step of embodiment 1 is utilized
(spectrum spectrum detects material in the range of 950-1650nm to the big aggressiveness content prediction calibrating patterns of Insoluble glutenin set up in 4
Absorbance, Testing index is Glutenin macropolymer content) sample is detected, and near infrared detection value (table 3) is obtained, detect
When measuring condition and being consistent during modeling.
The chemical score of 20 parts of samples is determined using chromatographiccondition identical with the step 2 of embodiment 1, as a result such as table 3.
Near-infrared prediction calibrating patterns testing result is compared with chemical score, 3 are as a result equally shown in Table.
The chromatography chemical score and model inspection value (AU/mg) of 3 20 parts of checking model flour samples of table
Note：Prediction standard deviation is 43.66AU/mg, and prediction related coefficient is 0.93.
As can be seen from Table 3, using the Glutenin macropolymer content detection model for the near infrared detection model determination set up
Enclose wider, prediction related coefficient reaches 0.93, and prediction standard deviation is 43.66AU/mg, and the model is in flour Insoluble glutenin
There is preferable application prospect in measure.
Claims (5)
1. a kind of near-infrared assay method of the big aggressiveness content of wheat flour Insoluble glutenin, it is characterised in that specific steps
It is as follows：
Prepare Glutenin macropolymer sample：
The big aggressiveness fluid sample of albumen is prepared using SDS- phosphate buffer extract solutions, it is standby；
Respectively step 1 is taken using nylon membrane syringe filters）The Glutenin macropolymer fluid sample of acquisition carries out chromatography, obtains
Obtain chemical assay result；
Step 1 is taken respectively）The mL of Glutenin macropolymer fluid test sample 1.0 ~ 1.2 of acquisition, gathers spectroscopic data；Spectrum is swept
Retouch 950 ~ 1650nm of interval, scanning times 10 times, temperature room temperature, resolution ratio 5nm；
Using deflected secondary air, to step 2）The chemical assay result and step 3 of acquisition）The spectroscopic data of acquisition, which is calculated, to be divided
Analysis, sets up forecast model；The coefficient of determination of the model is R2=0.89, the calibration standard deviation of model is RMSECV=36.40；
Using Unscrambler chemo metric softwares, the calibrating patterns of prediction are set up with reference to PLS method；
Spectrum baseline drift in measurement process is eliminated using the smooth Pretreated spectra modes of first derivative+S-G to predict setting up
The influence that causes of calibrating patterns, with the calibration standard deviation and the coefficient of determination R of institute's established model of validation-cross2It is pre- as evaluating
The major parameter of the calibrating patterns effect of survey；
Using with step 1）Identical method prepares the Glutenin macropolymer fluid sample of testing sample, utilizes step 5）Obtain
The calibrating patterns of prediction, and according to step 3）Methods described carries out near-infrared spectrum analysis to testing sample, that is, obtains and treat test sample
The content of the big aggressiveness of product Insoluble glutenin.
2. the near-infrared assay method of the big aggressiveness content of wheat flour Insoluble glutenin, its feature according to claim 1
It is, step 1）The utilization SDS- phosphate buffer extract solutions prepare the big aggressiveness fluid sample of albumen and referred to：Weigh respectively
20mg flour samples, add 1.6mL SDS- phosphate buffers extract solution concussion 10min, 17000g 5 min of centrifugation, abandon supernatant；
Add 1.6 mL SDS- phosphate buffer extract solutions again in precipitation, extract 20s using Ultrasonic cell smash, then
17000 g centrifuge 5 min, and the supernatant obtained is Glutenin macropolymer fluid sample；
The SDS- phosphate buffers extract solution refers to：The 0.5M phosphate buffers of the SDS containing mass volume ratio 0.5%, pH value
6.90。
3. the near-infrared assay method of the big aggressiveness content of wheat flour Insoluble glutenin, its feature according to claim 2
It is, step 1）The extracting parameter of the Ultrasonic cell smash is popped one's head in for 3mm, output work 10W.
4. the near-infrared assay method of the big aggressiveness content of wheat flour Insoluble glutenin, its feature according to claim 1
It is, step 2）The chromatography refers to, using high performance liquid chromatograph, and mobile phase is containing the trifluoroacetic acid of volume ratio 0.05%
50% acetonitrile solution, the mL/min of flow velocity 0.5.
5. according to the near-infrared assay method of the big aggressiveness content of one of the claim 1-4 wheat flour Insoluble glutenins,
Characterized in that, step 3）During the collection spectroscopic data, dress sample hardware is liquid quartz sample cell, and thickness is 1mm.
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