CN117250303A - Method for quantitatively determining binding force of rice starch grains and protein and components thereof - Google Patents
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- 235000013339 cereals Nutrition 0.000 title claims abstract description 69
- 102000004169 proteins and genes Human genes 0.000 title claims abstract description 51
- 108090000623 proteins and genes Proteins 0.000 title claims abstract description 51
- 238000000034 method Methods 0.000 title claims abstract description 14
- 229940100486 rice starch Drugs 0.000 title claims abstract description 14
- 229920002472 Starch Polymers 0.000 claims abstract description 78
- 235000019698 starch Nutrition 0.000 claims abstract description 78
- 239000008107 starch Substances 0.000 claims abstract description 78
- 235000007164 Oryza sativa Nutrition 0.000 claims abstract description 51
- 235000009566 rice Nutrition 0.000 claims abstract description 51
- 108010088751 Albumins Proteins 0.000 claims abstract description 18
- 102000009027 Albumins Human genes 0.000 claims abstract description 18
- 108010044091 Globulins Proteins 0.000 claims abstract description 18
- 102000006395 Globulins Human genes 0.000 claims abstract description 18
- 108010068370 Glutens Proteins 0.000 claims abstract description 18
- RWRDLPDLKQPQOW-UHFFFAOYSA-N Pyrrolidine Chemical compound C1CCNC1 RWRDLPDLKQPQOW-UHFFFAOYSA-N 0.000 claims abstract description 8
- 241000209094 Oryza Species 0.000 claims description 49
- 235000018102 proteins Nutrition 0.000 claims description 45
- 239000006228 supernatant Substances 0.000 claims description 34
- 235000013312 flour Nutrition 0.000 claims description 24
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 18
- 108060006613 prolamin Proteins 0.000 claims description 14
- 235000019624 protein content Nutrition 0.000 claims description 14
- 235000021312 gluten Nutrition 0.000 claims description 13
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 12
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 12
- 239000008187 granular material Substances 0.000 claims description 11
- 238000000926 separation method Methods 0.000 claims description 6
- 239000011780 sodium chloride Substances 0.000 claims description 6
- 238000005238 degreasing Methods 0.000 claims description 5
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 claims description 4
- 229910021642 ultra pure water Inorganic materials 0.000 claims description 4
- 239000012498 ultrapure water Substances 0.000 claims description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 4
- 238000009395 breeding Methods 0.000 claims description 3
- 230000001488 breeding effect Effects 0.000 claims description 3
- 235000013305 food Nutrition 0.000 abstract description 7
- 244000062793 Sorghum vulgare Species 0.000 abstract description 3
- 241000209140 Triticum Species 0.000 abstract description 3
- 235000021307 Triticum Nutrition 0.000 abstract description 3
- 240000008042 Zea mays Species 0.000 abstract description 3
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 abstract description 3
- 235000002017 Zea mays subsp mays Nutrition 0.000 abstract description 3
- 235000005822 corn Nutrition 0.000 abstract description 3
- 235000019713 millet Nutrition 0.000 abstract description 3
- 238000004458 analytical method Methods 0.000 abstract description 2
- 240000007594 Oryza sativa Species 0.000 abstract 2
- 238000005119 centrifugation Methods 0.000 description 6
- 238000007696 Kjeldahl method Methods 0.000 description 4
- 235000004252 protein component Nutrition 0.000 description 4
- 238000004737 colorimetric analysis Methods 0.000 description 3
- 239000002207 metabolite Substances 0.000 description 3
- 239000002245 particle Substances 0.000 description 3
- 235000008429 bread Nutrition 0.000 description 2
- 238000010411 cooking Methods 0.000 description 2
- 230000000875 corresponding effect Effects 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 238000006467 substitution reaction Methods 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
- 238000000576 coating method Methods 0.000 description 1
- 230000002596 correlated effect Effects 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
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- 238000012545 processing Methods 0.000 description 1
- 238000005215 recombination Methods 0.000 description 1
- 230000006798 recombination Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000004062 sedimentation Methods 0.000 description 1
- 238000011282 treatment Methods 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
- G01N31/00—Investigating or analysing non-biological materials by the use of the chemical methods specified in the subgroup; Apparatus specially adapted for such methods
- G01N31/002—Determining nitrogen by transformation into ammonia, e.g. KJELDAHL method
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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
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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
- G01N1/40—Concentrating samples
- G01N1/4055—Concentrating samples by solubility techniques
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- 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
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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
- G01N1/40—Concentrating samples
- G01N1/4055—Concentrating samples by solubility techniques
- G01N2001/4061—Solvent extraction
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Abstract
The invention relates to a method for quantitatively determining binding force of rice starch grains and proteins and components thereof in the technical field of agricultural product analysis, which sequentially determines binding force of albumin, globulin, glutelin and prolamine to starch grains, calculates average value to obtain binding force of starch grains and total proteins, and uses the value to evaluate or improve taste and texture of rice food, wherein the binding force of starch grains and proteins is higher as the C value is higher, and the viscosity and elasticity of starch grains are better. The method is not limited to measuring the binding force of starch grains and total proteins in rice, and can be used for measuring the binding force of starch grains and total proteins in other grains, such as wheat, corn, millet and the like.
Description
Technical Field
The invention relates to the technical field of agricultural product analysis, in particular to a technology for measuring the binding force of starch grains and protein of rice.
Background
Rice is used as a staple food for half of the population worldwide, and supplies basic energy for more than 60% of the population in China. Starch (90%, in the form of starch granules), protein (8%), which is the two most critical metabolites, accounts for about 98% of the dry weight of rice. The influence of the two on rice is not more than four aspects: metabolite content, composition, structure and binding force thereof. The former establishes quantitative description method of metabolite content, components and structure, lays foundation for the formation of rice yield and quality, and plays an important role in the development of rice industry. As the study goes further, the scholars further confirm that rice is closely related to the change of the binding force of the starch and protein: the protein can be tightly attached to the surface of starch body by competing with starch grains to bind water and disulfide bonds of the protein, so that excessive protein can be tightly attached to the surface of starch body, protein molecules are entangled with long-chain starch in the cooking process, the disintegration rate of starch structure is slowed down, the release of protons is inhibited, the viscosity of rice is reduced, and the taste of rice is deteriorated as a result, and the loose connection of starch and protein can greatly increase chalkiness, but also weaken the competition of two factors on bound water, so that researches on the taste quality and chalkiness of rice are obviously positively correlated. As described above, the binding force of starch grains and protein has a significant influence on the taste and quality of rice products. But there is currently a lack of methods to quantitatively describe the binding of starch particles to proteins and their components.
Disclosure of Invention
The invention aims to provide a method for quantitatively determining the binding force of rice starch grains and proteins and components thereof, wherein the binding force of the rice starch grains and total proteins is obtained through quantitative calculation so as to further evaluate or improve the taste and texture of rice food.
Therefore, the method for quantitatively determining the binding force of rice starch grains, protein and components thereof provided by the invention comprises the following steps:
step 1), determining the binding force of starch grains and albumin:
after degreasing rice flour, adding ultrapure water, oscillating uniformly, centrifuging by a centrifuge, and collecting supernatant to obtain albumin solution A1; adding water into the rest residue, centrifuging, separating supernatant, repeating for three times, and mixing the three repeated supernatants to obtain albumin solution B1; and respectively analyzing the protein content of A1 and B1 to obtain A2 and B2, wherein the binding force C1 of the starch granules and albumin is as follows: c1 (%) =b2/(a2+b2) 100;
step 2), determining the binding force of starch grains and globulin:
adding the residue at the bottom after the step 1) into NaCl solution, oscillating uniformly, centrifuging by a centrifuge, and collecting supernatant to obtain globulin solution A3; adding NaCl solution into the residue, centrifuging, separating supernatant, repeating for three times, and mixing the three repeated supernatants to obtain globulin solution B3; the protein contents of A3 and B3 are respectively analyzed to obtain A4 and B4, and the binding force C2 of starch grains and globulin is as follows: c2 (%) =b4/(a4+b4) 100;
step 3), determining the binding force of starch grains and gluten:
adding the residues at the bottom after the step 2) into NaOH solution, oscillating uniformly, centrifuging by a centrifuge, and collecting supernatant to obtain gluten solution A5; adding NaOH solution into the residue, centrifuging, separating supernatant, repeating for three times, and mixing the three repeated supernatants to obtain gluten solution B5; the protein contents of A5 and B5 are respectively analyzed to obtain A6 and B6, and the binding force C3 of starch grains and gluten is as follows: c3 (%) =b6/(a6+b6) 100;
step 4), determining the binding force of starch grains and prolamin:
adding the residue at the bottom after the step 3) into an ethanol solution, oscillating uniformly, centrifuging by a centrifuge, and collecting supernatant to obtain an prolamin solution A7; adding ethanol solution into the residue, centrifuging, separating supernatant, repeating for three times, and mixing the three repeated supernatants to obtain prolamin solution B7; and respectively analyzing the protein content of A7 and B7 to obtain A8 and B8, wherein the binding force C4 of the prolamin and the starch granules is as follows: c4 (%) =b8/(a8+b8) 100;
step 5), determining the binding force of starch grains and total protein:
the average value is calculated according to the formula c= (c1+c2+c3+c4)/4, wherein the C value in the formula represents the binding force of starch grains and total protein, the C value is used for evaluating or improving the taste and texture of rice food, the larger the C value is, the larger the binding force of starch grains and protein is, and the better the viscosity and elasticity of starch grains are.
Further, when rice flour is degreased, the rice flour is placed in a centrifuge tube, n-hexane is added for degreasing for a period of time, and then centrifugal separation is carried out, and supernatant is removed.
Further, steps 1) to 4) are carried out at a constant temperature of 20 ℃, the rotational speed of the centrifugal separation of albumin, globulin, gluten and prolamine is 3000 rpm, and the first separation time in the corresponding steps is 6min, 20min and 2min respectively; the corresponding time for centrifuging the residue in each step is 15min. Based on the difference of sedimentation coefficients of starch grains and different protein components in different solutions during centrifugation, the invention determines the centrifugation time under a certain rotation speed of the centrifuge, and can obtain the binding force of the starch grains and the total protein more accurately and rapidly under the time.
Compared with the prior art, the invention has the beneficial effects that: the rice flour is a combination of starch grains, protein and components thereof through adhesion, embedding, surrounding and the like. The starch grain and protein binding force is represented by coating the surface of the starch grain with a layer of protein, and the binding can make the starch grain more stable and prevent the starch grain from excessively decomposing or losing form during cooking or processing. At the same time, the existence of protein can also increase the viscosity and elasticity of starch grains and improve the taste and texture of food. For example, in bread making, starch granules combine with proteins in the dough to increase the elasticity and extensibility of the dough, making the bread softer and more elastic. The invention solves the problem that the binding force between starch grains and total protein in rice flour can not be quantitatively described. The binding force between albumin, globulin, glutelin and prolamin and starch granules is determined sequentially through the sequence of the steps, so that the mutual interference of different reagents obtained in detection on the detection effect is avoided, and the more accurate binding force between starch granules and total proteins is obtained. The taste and texture of rice food are evaluated or improved by the quantitative value, and the binding force of starch grains and protein is greater as the C value is greater, and the viscosity and elasticity of the starch grains are better. The method is not limited to measuring the binding force of starch grains and total proteins in rice, and can be used for measuring the binding force of starch grains and total proteins in other grains, such as wheat, corn, millet and the like.
Drawings
FIG. 1 is an electron microscope image of rice starch grains and combinations of starch grains and protein binding forces.
Wherein A is rice starch grains (protein-free state); b is the binding state of rice starch and protein (B type rice flour); c is the binding state of rice starch and protein (C-type rice flour).
Detailed Description
The invention will be further illustrated with reference to specific examples.
A method for quantitatively determining binding force of rice starch grains and proteins and components thereof is characterized by comprising the following steps:
(1) Determining the binding force of starch grains and albumin: 1g of rice flour is taken for degreasing, 4ml of ultrapure water is added for constant-temperature oscillation at 20 ℃ for 4h, then centrifugation is carried out at 3000 rpm for 6min, and supernatant fluid is collected to obtain albumin solution A1; the remaining residue was added with 4ml ultrapure water, centrifuged at 3000 rpm for 15min, repeated three times, and the supernatant of the three repetitions was combined to give albumin solution B1. The protein content in the A1 and B1 liquid is measured by adopting a Kjeldahl method or a colorimetric method and the like, and the protein content is A2 and B2 respectively; the binding force C1 between starch grains and albumin is as follows: c1 (%) =b2/(a2+b2) 100.
(2) Determining the binding force of starch grains and globulin: adding 4ml of 5% NaCl solution into the residue treated in the step (1), oscillating for 4 hours at 20 ℃, centrifuging at 3000 rpm for 6 minutes, and collecting supernatant to obtain globulin solution A3; the remaining residue was further added with 4ml of 5% NaCl solution, centrifuged at 3000 rpm for 15min, repeated three times, and the three repeated supernatants were combined to give globulin solution B3. Protein contents of A3 and B3 solutions, namely A4 and B4, are measured by adopting a Kjeldahl method or a colorimetric method and the like; the starch grain and globulin binding force C2 is: c2 (%) =b4/(a4+b4) 100.
(3) Determining the binding force of starch grains and gluten: adding 4ml of 0.02M NaOH solution (pH=11.0) into the residue treated in the step (2), shaking at 20 ℃ for 30min, centrifuging at 3000 rpm for 20min, and collecting supernatant to obtain gluten solution A5; the residue was further added to 4ml of 0.02m NaOH solution (ph=11.0), centrifuged at 3000 rpm for 15min, repeated 3 times, and the three repeated supernatants were combined to give gluten solution B5. Protein content of A5 and B5 solutions is measured by Kjeldahl method or colorimetry, etc., and is A6 and B6 respectively; the binding force C3 between starch grains and gluten is as follows: c3 (%) =b6/(a6+b6) 100.
(4) Determining the binding force of starch grains and prolamin: adding 3ml of 70% ethanol solution into the residue treated in the step (3), oscillating for 4h at 20 ℃, centrifuging at 3000 rpm for 2min, collecting supernatant to obtain prolamin solution A7, repeating the steps for 3 times from adding 3ml of 70% ethanol solution to centrifuging at 3000 rpm for 15min, and combining the three repeated supernatants to obtain prolamin solution B7. Protein content of A7 and B7 solutions, A8 and B8 respectively, was determined by Kjeldahl method, and binding force C4 between prolamin and starch granules was: c4 (%) =b8/(a8+b8) 100. Step 5), determining the binding force of starch grains and total protein:
(5) Calculating an average value according to a formula C= (C1+C2+C3+C4)/4, wherein a C value in the formula represents the binding force of starch grains and total protein, the C value is used for evaluating or improving the taste and texture of rice food in the cultivation and breeding fields, the larger the C value is, the larger the binding force of starch grains and protein is, and the better the viscosity and elasticity of starch grains are.
Table 1 is a table comparing the analytical forces of binding forces of starch and protein components of different types of rice under different centrifugation times. Rice flour samples with different protein contents and chalkiness are obtained from the same rice variety under different treatments, namely B-type rice flour and C-type rice flour (Table 1), wherein the B-type rice flour is high-protein low-chalkiness rice flour and the C-type rice flour is low-protein high-chalkiness rice flour, and the rice used for production is respectively from environment CO 2 Obtained by culturing in an atmosphere having a concentration of 400 and 580. Mu. Mol/mol, respectively, i.e., CO per mole of atmosphere 2 The rice which is obtained by growing in two environments with the amounts of 400 and 580 mu mol respectively is processed into rice flour, and the rice varieties are Nanjing 9108; the centrifuge (LXJ-IIB, shanghai Anting/Anke scientific instruments factory, shanghai, china) has a rotational speed of 3000 rpm. Binding force of two types of rice flour starch and protein and components is shown in table 1; combining the difference and ratio of binding force of two types of rice flour, albumin, globulin, glutelin and prolamin are respectively 3000 rpm centrifugation for 6min, 20min and 2min was the best discrimination. The results in Table 1 show that the binding force of the starch granules of the C-type rice flour with albumin, globulin, glutelin, prolamine and total protein is 2.21, 3.95, 23.58, 9.29 and 9.76 percent higher than that of the B-type rice flour, respectively, and the binding force of the starch granules of the low-protein and chalky C-type rice flour with protein and protein components is larger. The greater the C value, the greater the binding of the starch particles to the protein and the better the viscosity and elasticity of the starch particles. The C value can be used for guiding cultivation and breeding and evaluating or improving the taste and texture of rice.
TABLE 1 analytical force comparison of binding force of starch and protein Components for different types of Rice at different centrifugation times
The invention is not limited to the above embodiments, and based on the technical solution disclosed in the invention, a person skilled in the art may make some substitutions and modifications to some technical features thereof, such as recombination of rotational speed of the centrifuge and centrifugal time, without creative effort, according to the technical disclosure, and these substitutions and modifications are all within the scope of the invention. The invention is not limited to rice, but can be used for other grains such as wheat, corn, millet, etc.
Claims (3)
1. A method for quantitatively determining binding force of rice starch grains and proteins and components thereof is characterized by comprising the following steps:
step 1), determining the binding force of starch grains and albumin:
after degreasing rice flour, adding ultrapure water, oscillating uniformly, centrifuging by a centrifuge, and collecting supernatant to obtain albumin solution A1; adding water into the rest residue, centrifuging, separating supernatant, repeating for three times, and mixing the three repeated supernatants to obtain albumin solution B1; and respectively analyzing the protein content of A1 and B1 to obtain A2 and B2, wherein the binding force C1 of the starch granules and albumin is as follows: c1 (%) =b2/(a2+b2) 100;
step 2), determining the binding force of starch grains and globulin:
adding the residue at the bottom after the step 1) into NaCl solution, oscillating uniformly, centrifuging by a centrifuge, and collecting supernatant to obtain globulin solution A3; adding NaCl solution into the residue, centrifuging, separating supernatant, repeating for three times, and mixing the three repeated supernatants to obtain globulin solution B3; the protein contents of A3 and B3 are respectively analyzed to obtain A4 and B4, and the binding force C2 of starch grains and globulin is as follows: c2 (%) =b4/(a4+b4) 100;
step 3), determining the binding force of starch grains and gluten:
adding the residues at the bottom after the step 2) into NaOH solution, oscillating uniformly, centrifuging by a centrifuge, and collecting supernatant to obtain gluten solution A5; adding NaOH solution into the residue, centrifuging, separating supernatant, repeating for three times, and mixing the three repeated supernatants to obtain gluten solution B5; the protein contents of A5 and B5 are respectively analyzed to obtain A6 and B6, and the binding force C3 of starch grains and gluten is as follows: c3 (%) =b6/(a6+b6) 100;
step 4), determining the binding force of starch grains and prolamin:
adding the residue at the bottom after the step 3) into an ethanol solution, oscillating uniformly, centrifuging by a centrifuge, and collecting supernatant to obtain an prolamin solution A7; adding ethanol solution into the residue, centrifuging, separating supernatant, repeating for three times, and mixing the three repeated supernatants to obtain prolamin solution B7; and respectively analyzing the protein content of A7 and B7 to obtain A8 and B8, wherein the binding force C4 of the prolamin and the starch granules is as follows: c4 (%) =b8/(a8+b8) 100;
step 5), determining the binding force of starch grains and total protein:
calculating an average value according to a formula C= (C1+C2+C3+C4)/4, wherein a C value in the formula represents the binding force of starch grains and total protein, the C value is used for evaluating or improving the taste and texture of rice in the cultivation and breeding fields, the larger the C value is, the larger the binding force of starch grains and protein is, and the better the viscosity and elasticity of starch grains are.
2. The method for quantitatively determining binding force of rice starch grains to proteins and components thereof according to claim 1, wherein: when the rice flour is degreased, the rice flour is placed in a centrifuge tube, n-hexane is added for degreasing for a period of time, and then centrifugal separation is carried out, and the supernatant is removed.
3. The method for quantitatively determining binding force of rice starch grains to proteins and components thereof according to claim 1, wherein: steps 1) to 4) are carried out at a constant temperature of 20 ℃, the rotational speed of the centrifugal separation of albumin, globulin, glutelin and prolamine is 3000 rpm, and the first separation time in the corresponding steps is 6min, 20min and 2min respectively; the corresponding time for centrifuging the residue in each step is 15min.
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