CN104006998A - Method for separating different components of starch of rice and determining contents thereof - Google Patents
Method for separating different components of starch of rice and determining contents thereof Download PDFInfo
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- CN104006998A CN104006998A CN201410256050.6A CN201410256050A CN104006998A CN 104006998 A CN104006998 A CN 104006998A CN 201410256050 A CN201410256050 A CN 201410256050A CN 104006998 A CN104006998 A CN 104006998A
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Abstract
The invention discloses a method for separating different components of starch of rice and determining the contents thereof. The method comprises the following steps: taking more than 10 rice grains randomly, separating the different components of the starch and determining the contents of the different components of the starch of all the rice grains respectively; adding 10ml of pure water into the rice grains, and gelatinizing in water bath at the temperature of 80 DEG C, thereby obtaining a starch turbid liquid; carrying out centrifugation at the temperature of 3 DEG C for 30 minutes by using relative centrifugal force of 20000g, thereby obtaining a supernatant and a precipitate; taking out the supernatant as a to-be-detected sample, adding 1.5mg/ml of KOH solution by 10ml, then carrying out centrifugal separation, and taking out the supernatant as the other to-be-detected sample; increasing the concentration of KOH solution added into the precipitate every time by a 1.5mg/ml gradient, carrying out centrifugal separation on the precipitate, and repeatedly such operations till the obtained precipitate can not be dissolved, thereby obtaining a series of samples to be determined; then respectively carrying out determination on the content of apparent amylose on the to-be-detected samples according to the national standard GB/T15683-200. The method disclosed by the invention has the advantages that the content of the apparent amylose of different components of the starch in the rice of rice breeding materials can be determined, and basis is provided for the breeding work of new-variety rice with good mouthfeel and quality.
Description
Technical field
The invention belongs to rice quality assay method technical field, be specifically related to a kind of for the different separation of each component of starch of the rice degree of polymerization and the method for assay thereof.
Background technology
Press the mouthfeel quality of rice, rice varieties can be divided into four large classes substantially: common rice variety, common japonica rice variety, soft rice kind, glutinous rice variety, the rice of this four types rice varieties exists very big-difference in mouthfeel quality.In order to improve the quality of rice varieties, meet the different demands of people to rice mouthfeel quality, breeder need study affecting the factor of rice mouthfeel quality.People find already, exist larger difference, Japanese scholars and international paddy rice to cooperate also to find the vital role of amylopectin long-chain in determining Texture of Cooked between the kind of similar amylose content on Texture of Cooked.Now clear and definite, the true cause that causes Texture of Cooked difference is not content of starch (amylose content, AC), but the number of long-chain and length (degree of polymerization) size of chain on amylopectin.
1958, between the discoveries such as Williams " iodine dyes value " and content of starch (amylose content, AC), there is correlativity, proposed with colorimetric method for determining rice amylose assay method.Afterwards, improved the method became general international standard.But, between the AC that this what is called " AC " of being calculated by " iodine dyes value " and rice really have, there is certain difference.Because the amylopectin of part long-chain can react with iodine equally.So Takeda equals the AC value recording with iodine colorimetry to be called to apparent amylose content in 1987.From forming, in fact apparent amylose comprises that two parts form, i.e. the long-chain of real amylose and part amylopectin.That is to say, the long-chain of real amylose and part amylopectin all can react with iodine reagent (developer) and develop the color, everyly can all can observe intuitively color with the starch of iodine reagent (developer) combination, thereby, the absorbance that can pass through the solution of its colour developing of spectrophotometric determination, is just called apparent amylose.
Amylose or amylopectin are all the glucosides being polymerized through hydroxyl dehydration by a plurality of glucose molecules.Rice starch is comprised of the different glucoside chain of the degree of polymerization, contains great amount of hydroxy group.Because hydroxyl in starch exists with hydrogen bond state, intermolecular have a huge aggregation force, stops hydrone to the infiltration of starch granules and solvation; Meanwhile, also have different branch degree amyloses and amylopectin, winding intertwines, and causes rice starch water-soluble hardly.Therefore,, in order to improve starch dissolution performance, must destroy the retrogradation phenomenon that hydrogen bond causes.
The common method of destroying starch glucose interchain hydrogen bond comprises starch fluid body high-temperature heating, adds the reagent such as NaOH or potassium hydroxide.
The method key step that prior art is measured rice starch content is that rice is clayed into power, and adds certain density sig water to dissolve the rear absorbance that detects coloured moiety, can only calculate the part content of apparent amylose.This kind of method can not demonstrate the content of apparent amylose in the different component that in starch, the degree of polymerization is different with starch branch degree, thereby can not reflect better the apparent amylose content of the mouthfeel situation that affects.The defect of prior art and deficiency can not meet the needs to rice starch different component assay.
Summary of the invention
The technical problem to be solved in the present invention is to overcome the part content that calculates apparent amylose that prior art existence can only be general, content and the distribution situation thereof that can not reflect the apparent amylose of the different component that the degree of polymerization is different, so that the starch ingredients that can not reflect better the rice varieties of different mouthfeel quality, affects the screening that paddy rice is improved the breeding material of taste quality.
The object of the invention is, by directly measuring affecting the content of apparent amylose in the different component that in the rice of taste quality, the degree of polymerization is different, provides foundation for improving the breeding material screening operation of paddy rice taste quality.Therefore the invention provides the method for the separation of a kind of rice starch different component and assay.
The inventive method is to separate being wound around different polymerization degree mixed in together and the starch of branch degree, so that the content of the apparent amylose of the different component that in separation determination starch, the degree of polymerization is different.
The technical scheme of the method for a kind of rice starch different component provided by the present invention separation and assay comprises the following steps:
(1) get at random 10 above complete grain of rices of removing clever shell, planting skin and embryo, each grain of rice is operation as follows all:
A. weigh and add the gelatinization 1 hour in 80 ℃ of water-baths of 10ml pure water after a grain of rice, be cooled to room temperature and obtain starch suspension;
B. starch suspension is obtained to supernatant and precipitation by centrifugal condition centrifuging, supernatant is for treating test sample, mark 0; In this precipitation, add 1.5mg/ml KOH solution 10ml, after shaking up, give over to centrifuging next time; Described centrifugal condition is: relative centrifugal force(RCF) 20000g, 4 ℃ of centrifugal 30min;
C. the precipitation and centrifugal separation of KOH solution will have been added in step (1) B, centrifugal condition is: relative centrifugal force(RCF) 20000g, 4 ℃ of centrifugal 30min, obtain supernatant and precipitation, using the supernatant of gained as treating test sample, be labeled as in order 1, the precipitation of gained is added to the centrifuging again of KOH solution again, and so repeated centrifugation lock out operation, can not dissolve until gained precipitates, each centrifugal condition is: relative centrifugal force(RCF) 20000g, 4 ℃ of centrifugal 30min; The KOH solution at every turn gained precipitation being added is 10ml, but the concentration of the KOH solution adding increases progressively with 1.5mg/ml gradient, and the supernatant of each centrifuging gained is treats test sample mark in order, obtains a series of test samples for the treatment of;
(2) by the mensuration of State Standard of the People's Republic of China GB/T 15683-2008 Rice Amylose content, to what obtain in step (1) C, a series ofly treat that test sample carries out respectively the mensuration of amylose content.
The present invention on to the basis of different centrifugal force (10000g, 20000g, 40000g) and centrifugation time (10min, 20min, 30min) contrast test, draws: relative centrifugal force(RCF) is not enough and the time falls short of all may cause precipitating and supernatant cannot be completely separated.Through repetition test, relative centrifugal force(RCF) 20000g, centrifugation time 30min, temperature is 4 degree, effect is best.
Compared with prior art, the invention has the beneficial effects as follows:
The present invention by starch repeatedly centrifuging until precipitation can not dissolve again, and suitable parameter (concentration gradient of centrifugal condition and the concentration of KOH solution) reaches the different different component continuous separated effect from low polymerization degree to high polymerization degree of the degree of polymerization in starch, can effectively determine content and the distribution situation thereof of the apparent amylose of starch different component in the rice of different rice varieties of taste quality difference (taste quality differs greatly), therefore, adopt the inventive method can measure content and the distribution situation (distribution audio-visual picture) thereof of the apparent amylose of starch different component in rice breed, to paddy rice taste quality improvement seed selection new varieties, work provides direct foundation.
Accompanying drawing explanation
Fig. 1 is relative apparent amylose content distribution audio-visual picture in rice variety 93-11 rice starch different component.In figure horizontal ordinate be KOH solution concentration 1.5 mg/ml, 3 mg/ml ... 13.5mg/ml, ordinate is relative apparent amylose content.
Fig. 2 is relative apparent amylose content distribution audio-visual picture in the extensive 290 rice starch different components of soft rice kind cloud.In figure horizontal ordinate be KOH solution concentration 1.5 mg/ml, 3 mg/ml ... 13.5mg/ml, ordinate is relative apparent amylose content.
Fig. 3 is relative apparent amylose content distribution audio-visual picture in the fine rice starch different component of japonica rice Japan.In figure horizontal ordinate be KOH solution concentration 1.5 mg/ml, 3 mg/ml ... 13.5mg/ml, ordinate is relative apparent amylose content.
Fig. 4 is relative apparent amylose content distribution audio-visual picture in the glutinous No. 1 rice starch different component of glutinous rice variety literary composition.In figure horizontal ordinate be KOH solution concentration 1.5 mg/ml, 3 mg/ml ... 13.5mg/ml, ordinate is relative apparent amylose content.
Embodiment
The invention will be further described below to use four examples of implementation.
Four rice taste quality rice varieties that there were significant differences of generally acknowledging in embodiment, have been adopted: rice variety 93-11, soft rice kind cloud are extensive 290, japonica rice Japan is fine, glutinous rice variety civilian glutinous No. 1.These four rice varieties and rice thereof are commercially available.
Embodiment 1 carries out the mensuration of amylose content, the namely mensuration of apparent amylose content to the grain of rice of rice variety 93-11 rice.
Embodiment 2 is to the soft rice of the soft rice extensive 290(of kind cloud Yunnan high-quality) grain of rice of rice carries out the mensuration of amylose content, the mensuration of apparent amylose content namely.
Embodiment 3 carries out the mensuration of amylose content, the namely mensuration of apparent amylose content to the grain of rice of the fine rice of japonica rice Japan.
Embodiment 4 carries out the mensuration of amylose content, the namely mensuration of apparent amylose content to the grain of rice of glutinous No. 1 rice of glutinous rice variety literary composition.
The rice starch different component separation of above four embodiment and the method for assay are all as follows:
(1) get at random 10 above complete grain of rices of removing clever shell, planting skin and embryo, each grain of rice is operation as follows all:
A. a grain of rice adds the gelatinization 1 hour in 80 ℃ of water-baths of 10ml pure water after weighing, and is cooled to room temperature and obtains starch suspension;
B. starch suspension is obtained to supernatant and precipitation by centrifugal condition centrifuging, supernatant is for treating test sample, mark 0; In this precipitation, add 1.5mg/ml KOH solution 10ml, after shaking up, give over to centrifuging next time; Described centrifugal condition is: relative centrifugal force(RCF) 20000g, 4 ℃ of centrifugal 30min;
C. in step (1) B, added the precipitation and centrifugal separation of 1.5mg/ml KOH solution, using the supernatant of gained as treating test sample, be labeled as in order 1, add again KOH solution centrifugal separated the precipitation of gained, like this repeated centrifugation lock out operation (precipitation each centrifuging being obtained add KOH solution after centrifuging again), until gained precipitation can not be dissolved, each centrifugal condition is: relative centrifugal force(RCF) 20000g, 4 ℃ of centrifugal 30min; The KOH solution at every turn gained precipitation being added is 10ml, but the concentration of the KOH solution adding increases progressively with 1.5mg/ml gradient, as 1.5 mg/ml, 3 mg/ml ... 1.35mg/ml.The supernatant of each centrifuging gained is treats test sample mark in order, obtains a series of test samples for the treatment of;
(2) by the mensuration of State Standard of the People's Republic of China GB/T 15683-2008 Rice Amylose content, to what obtain in step (1) C, a series ofly treat that test sample carries out respectively the mensuration of amylose content (the mensuration of amylose content be the mensuration of apparent amylose content).
The rice starch of the rice varieties of different Cooking Qualities is starch solution and the iodine chrominance response with different polymerization degree and the branch degree of the separation of 1.5mg/ml concentration gradient incremental manner with KOH solution, with spectrophotometric instrumentation absorbance A, according to the apparent amylose content of typical curve calculation of starch different component.And with the relative apparent amylose content of following formula calculation of starch different component, the results are shown in Table 1.
Computing formula:
Apparent amylose content=single component apparent amylose quality/mono-grain of rice gross mass
The relative total apparent amylose content of apparent amylose content=single component apparent amylose content/mono-grain of rice
According to the data of table 1, take KOH concentration as horizontal ordinate, apparent amylose content is ordinate relatively, sees accompanying drawing 1, Fig. 2, Fig. 3, Fig. 4.
Result shows: according to peak type, the different rice varieties of taste quality obviously can be distinguished.The rice variety (as 93-11) of common rice matter is bimodal pattern, and soft rice kind (cloudlike extensive 290) also has bimodal, but postpeak is wider and short; The japonica rice variety of common quality (as fine in Japan) is 3 peak types, and glutinous rice variety (as glutinous in literary composition No. 1) is single peak type.
In collection of illustrative plates, the ordinate of every duplicate samples is relative apparent amylose content number percent, is representing that single component apparent amylose content accounts for the number percent of the total apparent amylose content of single grain of rice.The difference of peak value also can reflect interracial difference, and this separation and content assaying method can demonstrate starch component feature between different cultivars comparatively intuitively and accurately.
The method can apply in other rice varieties, also can expand sampling grain of rice number, increases individual sample whole representative.Use this disposal route can summarize the feature of a certain kind or the comprehensive starch component of the sample of a certain batch.
By embodiment, can find out that the method can distinguish the difference between different rice varieties, and obtain overall target by statistical method.The breeding of new variety work good for breeding rice taste quality provides direct foundation.
Claims (1)
1. a method for the separation of rice starch different component and assay, is characterized in that comprising the following steps:
(1) get at random 10 above complete grain of rices of removing clever shell, planting skin and embryo, each grain of rice is operation as follows all:
A. weigh and add the gelatinization 1 hour in 80 ℃ of water-baths of 10ml pure water after a grain of rice, be cooled to room temperature and obtain starch suspension;
B. starch suspension is obtained to supernatant and precipitation by centrifugal condition centrifuging, supernatant is for treating test sample, mark 0; In this precipitation, add 1.5mg/ml KOH solution 10ml, after shaking up, give over to centrifuging next time; Described centrifugal condition is: relative centrifugal force(RCF) 20000g, 4 ℃ of centrifugal 30min;
C. the precipitation and centrifugal separation of KOH solution will have been added in step (1) B, centrifugal condition is: relative centrifugal force(RCF) 20000g, 4 ℃ of centrifugal 30min, obtain supernatant and precipitation, using the supernatant of gained as treating test sample, be labeled as in order 1, the precipitation of gained is added to the centrifuging again of KOH solution again, and so repeated centrifugation lock out operation, can not dissolve until gained precipitates, each centrifugal condition is: relative centrifugal force(RCF) 20000g, 4 ℃ of centrifugal 30min; The KOH solution at every turn gained precipitation being added is 10ml, but the concentration of the KOH solution adding increases progressively with 1.5mg/ml gradient, and the supernatant of each centrifuging gained is treats test sample mark in order, obtains a series of test samples for the treatment of;
(2) by the mensuration of State Standard of the People's Republic of China GB/T 15683-2008 Rice Amylose content, to what obtain in step (1) C, a series ofly treat that test sample carries out respectively the mensuration of amylose content.
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Cited By (2)
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| CN104833671A (en) * | 2015-01-09 | 2015-08-12 | 中国水稻研究所 | Measurement method of absolute amylase content of rice |
| CN110261332A (en) * | 2019-05-30 | 2019-09-20 | 扬州大学 | A kind of method of simple grain rice measurement content of amylose in rice |
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Cited By (3)
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|---|---|---|---|---|
| CN104833671A (en) * | 2015-01-09 | 2015-08-12 | 中国水稻研究所 | Measurement method of absolute amylase content of rice |
| CN104833671B (en) * | 2015-01-09 | 2017-07-21 | 中国水稻研究所 | A kind of assay method of the absolute amylose content of rice |
| CN110261332A (en) * | 2019-05-30 | 2019-09-20 | 扬州大学 | A kind of method of simple grain rice measurement content of amylose in rice |
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