CN117186264A - Green and efficient highland barley beta-glucan extraction method - Google Patents
Green and efficient highland barley beta-glucan extraction method Download PDFInfo
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- CN117186264A CN117186264A CN202311304991.8A CN202311304991A CN117186264A CN 117186264 A CN117186264 A CN 117186264A CN 202311304991 A CN202311304991 A CN 202311304991A CN 117186264 A CN117186264 A CN 117186264A
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- 235000007340 Hordeum vulgare Nutrition 0.000 title claims abstract description 34
- FYGDTMLNYKFZSV-URKRLVJHSA-N (2s,3r,4s,5s,6r)-2-[(2r,4r,5r,6s)-4,5-dihydroxy-2-(hydroxymethyl)-6-[(2r,4r,5r,6s)-4,5,6-trihydroxy-2-(hydroxymethyl)oxan-3-yl]oxyoxan-3-yl]oxy-6-(hydroxymethyl)oxane-3,4,5-triol Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1OC1[C@@H](CO)O[C@@H](OC2[C@H](O[C@H](O)[C@H](O)[C@H]2O)CO)[C@H](O)[C@H]1O FYGDTMLNYKFZSV-URKRLVJHSA-N 0.000 title claims abstract description 30
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- 238000000034 method Methods 0.000 claims abstract description 34
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- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 21
- 108090000790 Enzymes Proteins 0.000 claims abstract description 15
- 102000004190 Enzymes Human genes 0.000 claims abstract description 15
- 229940088598 enzyme Drugs 0.000 claims abstract description 15
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- 102000004139 alpha-Amylases Human genes 0.000 claims abstract description 9
- 108090000637 alpha-Amylases Proteins 0.000 claims abstract description 9
- 229940024171 alpha-amylase Drugs 0.000 claims abstract description 9
- 108010019160 Pancreatin Proteins 0.000 claims abstract description 6
- 239000000284 extract Substances 0.000 claims abstract description 6
- 229940055695 pancreatin Drugs 0.000 claims abstract description 6
- 239000012153 distilled water Substances 0.000 claims abstract description 5
- 238000000227 grinding Methods 0.000 claims abstract description 5
- 238000002386 leaching Methods 0.000 claims abstract description 4
- 238000005406 washing Methods 0.000 claims abstract description 4
- 210000000582 semen Anatomy 0.000 claims abstract description 3
- 238000004108 freeze drying Methods 0.000 claims abstract 2
- 230000000415 inactivating effect Effects 0.000 claims abstract 2
- 238000002156 mixing Methods 0.000 claims abstract 2
- 238000000643 oven drying Methods 0.000 claims abstract 2
- 238000001556 precipitation Methods 0.000 claims abstract 2
- 239000002244 precipitate Substances 0.000 claims description 8
- 238000005119 centrifugation Methods 0.000 claims description 5
- 238000010992 reflux Methods 0.000 claims description 3
- 238000002481 ethanol extraction Methods 0.000 claims description 2
- 239000007788 liquid Substances 0.000 claims description 2
- 238000012869 ethanol precipitation Methods 0.000 claims 1
- 239000012535 impurity Substances 0.000 abstract description 3
- 239000000126 substance Substances 0.000 abstract description 3
- 230000009849 deactivation Effects 0.000 abstract 1
- 238000001914 filtration Methods 0.000 abstract 1
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 9
- 235000019441 ethanol Nutrition 0.000 description 8
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- 238000003809 water extraction Methods 0.000 description 5
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 3
- 238000001816 cooling Methods 0.000 description 3
- 235000013312 flour Nutrition 0.000 description 3
- 230000002779 inactivation Effects 0.000 description 3
- 206010009944 Colon cancer Diseases 0.000 description 2
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N Sodium azide Chemical compound [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 description 2
- 239000008280 blood Substances 0.000 description 2
- 210000004369 blood Anatomy 0.000 description 2
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 description 2
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- VWDWKYIASSYTQR-UHFFFAOYSA-N sodium nitrate Chemical compound [Na+].[O-][N+]([O-])=O VWDWKYIASSYTQR-UHFFFAOYSA-N 0.000 description 2
- 238000000967 suction filtration Methods 0.000 description 2
- 238000009777 vacuum freeze-drying Methods 0.000 description 2
- RWSOTUBLDIXVET-UHFFFAOYSA-N Dihydrogen sulfide Chemical compound S RWSOTUBLDIXVET-UHFFFAOYSA-N 0.000 description 1
- KDXKERNSBIXSRK-UHFFFAOYSA-N Lysine Natural products NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 description 1
- 239000004472 Lysine Substances 0.000 description 1
- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
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Abstract
The application discloses a green and efficient highland barley beta-glucan extracting method, which comprises the following steps: grinding highland barley into powder; grinding semen Avenae Nudae to obtain powder; enzyme deactivation and impurity removal of ethanol: extracting highland barley powder with 80% ethanol, filtering and oven drying; leaching with hot water: extracting with distilled water, centrifuging, and mixing the supernatants; enzyme removal: sequentially adding high temperature resistant alpha-amylase, saccharifying enzyme and pancreatin into the supernatant for enzymolysis, inactivating enzyme, centrifuging, and concentrating the supernatant; crude extraction: after alcohol precipitation, centrifugal washing, adding water and freeze-drying to obtain the beta-glucan extract. The application mainly adopts clean energy sources such as ethanol, water and the like, and does not use organic chemical substances; the method is simple to operate, low in equipment cost, and energy-saving and emission-reducing; the extraction efficiency is higher, and the industrial implementation is easy.
Description
Technical Field
The application relates to the field of highland barley functional components, in particular to a green and efficient highland barley beta-glucan extracting method.
Background
The highland barley in Qinghai province of China has wide planting area, high yield and excellent quality, but the highland barley industry has few deep processing enterprises, small scale, low technological content, low grade, poor quality, lagging processing technology and low added value of products, and cannot drive the development of highland barley production bases. Highland barley is rich in nutrition, 18 amino acids, starch and multiple vitamins necessary for human body, and the protein content and lysine content are higher than those of wheat and rice.
The beta-glucan plays a vital role in controlling blood sugar level, reducing blood fat, reducing cholesterol level, enhancing immunity and the like, can prevent or assist in improving type II diabetes and colon cancer, can improve serum short chain fatty acid level, and promotes lipid metabolism of liver. Can also be used as raw material in food industry. In addition, highland barley beta-glucan shows very high digestion resistance and can overcome alpha-amylase hydrolysis and gastric acid of human. Can reduce specific intestinal flora and hydrogen sulfide generation, thereby reducing risk of colon cancer generation and having obvious anti-fatigue effect. The method for extracting beta-glucan mainly comprises a three-phase distribution method, an ultrasonic-microwave synergistic method, an ultrasonic auxiliary extraction, a Microwave Auxiliary (MAE) extraction, a hot water extraction method, an ultrasonic method, a microwave method, a water extraction and alkali extraction method, an ultrahigh pressure and ultrasonic method, an acid hydrolysis method, an alkaline extraction method, a bioconversion method or a microbial fermentation method, a pressurized hot water extraction method, and the like, and is purified by an ammonium sulfate precipitation method, a column separation method, dialysis and the like. Although the methods are various, the extraction rate is low, the process is complex, the steps are complicated, the energy consumption is high, and the method is difficult to realize in the industrial application process.
Disclosure of Invention
The application discloses a green and efficient highland barley beta-glucan extraction method, which aims at the potential problems of reaction in documents and practice, adopts a hot water extraction method which is energy-saving and beneficial to industrial application, optimizes the extraction process steps, refines the step parameters, and further improves the extraction efficiency so as to meet the industrial extraction requirement.
The technical scheme of the application is as follows:
a green and efficient method for extracting highland barley beta-glucan comprises the following steps:
1. highland barley flour mill
Grinding semen Avenae Nudae to obtain barley flour. Screening through a 100 mesh screen in a laboratory mill has shown that the size of the abrasive particles affects the extraction efficiency, so that in the energy-saving process, an optimal abrasive particle size is selected as much as possible, and the beta-glucan is dissolved as much as possible in the extraction process.
2. Reflux extraction of ethanol
Highland barley powder and 80% ethanol are mixed according to the following ratio of 1:10w/v, and reflux-extracting at 80℃for 2h. After suction filtration, the precipitate is baked to constant weight at 60 ℃, and is sieved to 100 meshes after being crushed by a pulverizer.
3. Leaching with hot water
The ground highland barley powder is extracted with distilled water at 52 ℃ for 2h (feed-liquid ratio is 1:10 w/v), and then centrifuged at 5000rpm for 10min. This step was repeated 3 times and the supernatants were combined.
4. Enzyme impurity removal
10U/g high temperature resistant alpha-amylase is added into the supernatant, the pH is kept at 6.5, the mixture is heated for 2 hours in a water bath at 95 ℃, and the mixture is cooled to 60 ℃. pH4.5 was adjusted with dilute hydrochloric acid, 200U/g of saccharifying enzyme was added, and the reaction was carried out at 60℃for 30 minutes. The pH was adjusted to 7.5 with 0.1mol/L sodium hydroxide solution, and 200U/mg pancreatin was added thereto for reaction at 50℃for 1 hour. And (3) placing the mixture in a water bath at 100 ℃ for inactivation for 20 min after enzymolysis, and cooling to room temperature. The precipitate was removed by centrifugation at 5000rpm for 15min and the supernatant was concentrated.
5. Obtaining crude extract
Adding absolute ethanol to 80%, precipitating with ethanol at 5000rpm, centrifuging for 15min, washing the precipitate with absolute ethanol twice, dissolving in water, vacuum freeze drying to remove ethanol, and lyophilizing to obtain water extract beta-glucan extract (HBBG).
Advantageous effects
The method for extracting the beta-glucan changes the combination of enzymes and reduces the use amount of the enzymes as a whole. Compared with other physical or chemical extraction methods, the method mainly adopts clean energy sources such as ethanol, water and the like, and does not use organic chemical substances; the method is simple to operate, low in equipment cost, and energy-saving and emission-reducing; the extraction efficiency is higher, and the industrial implementation is easy.
Detailed Description
The following describes in detail the examples of the present application, which are implemented on the premise of the technical solution of the present application, and detailed embodiments and specific operation procedures are given, but the scope of protection of the present application is not limited to the following examples.
Example 1
A preparation method for extracting highland barley beta-glucan comprises the following steps:
(1) Highland barley flour mill
Pulverizing 50g of whole highland barley grains with a pulverizer, sieving with 100 mesh sieve, and keeping at 4deg.C.
(2) Ethanol extraction
50g of highland barley powder is soaked in 500ml of 80% ethanol and extracted for 2 hours under reflux at 80 ℃. After suction filtration, the precipitate is baked to constant weight at 60 ℃, and is sieved to 100 meshes after being crushed by a pulverizer.
(3) Leaching with hot water
The ground highland barley powder was extracted with distilled water at 52℃for 2h, then centrifuged at 5000rpm for 10min. This step was repeated 3 times and the supernatants were combined.
(4) Enzyme impurity removal
10U/g high temperature resistant alpha-amylase is added into the supernatant, the pH is kept at 6.5, the mixture is heated for 2 hours in a water bath at 95 ℃, and the mixture is cooled to 60 ℃. pH4.5 was adjusted with 1mol/L dilute hydrochloric acid, 200U/g of saccharifying enzyme was added, and the reaction was carried out at 60℃for 30 minutes. The pH was adjusted to 7.5 with 2mol/L sodium hydroxide solution, and 200U/mg pancreatin was added thereto for reaction at 50℃for 1 hour. And (3) placing the mixture in a water bath at 100 ℃ for inactivation for 20 min after enzymolysis, and cooling to room temperature. The precipitate was removed by centrifugation at 5000rpm for 15min and the supernatant was concentrated.
(5) Obtaining crude extract
Adding absolute ethyl alcohol to 80%, precipitating with ethanol, centrifuging at 5000rpm for 15min, washing precipitate with absolute ethyl alcohol twice, dissolving again with water, vacuum freeze drying to remove ethanol, and lyophilizing to obtain water extract beta-glucan extract (HBBG).
Comparative example 1
The difference from example 1 is that step (2) is omitted and the other steps are the same, and beta-glucan is obtained by extraction.
Comparative example 2
The difference from example 1 is that the hot water extraction in step (3) was performed for 2 hours without repeating the extraction three times.
Comparative example 3
The difference from example 1 is that the high temperature resistant alpha-amylase in step (4) is replaced by alpha-amylase, and the other steps are the same, and the beta-glucan is obtained by extraction.
Comparative example 4
The difference from example 1 is that 10U/g of high temperature resistant alpha-amylase was added to the supernatant in step (4), the pH was kept at 6.5, heated in a 95℃water bath for 2 hours, and cooled to 60 ℃. The pH was adjusted to 7.5 with 2mol/L sodium hydroxide solution, and 200U/mg pancreatin was added thereto for reaction at 50℃for 1 hour. pH4.5 was adjusted with 1mol/L dilute hydrochloric acid, 200U/g of saccharifying enzyme was added, and the reaction was carried out at 60℃for 30 minutes. And (3) placing the mixture in a water bath at 100 ℃ for inactivation for 20 min after enzymolysis, and cooling to room temperature. The precipitate was removed by centrifugation at 5000rpm for 15min and the supernatant was concentrated.
The other steps are the same.
Comparative example 5
The conditions of comparative examples 1 to 3 were combined and the other conditions were unchanged, and beta glucan was obtained by extraction.
The extracted beta-glucan of example 1 and comparative examples 1 to 5 was examined to obtain purity and extraction yield.
Testing of purity: the purity of beta-glucan was measured according to the method of AOAC (995.16) using a commercial detection kit (Megazyme International Ireland, bray, ireland).
And (3) detection of extraction rate:
determination of the molecular weight: the (polydispersity) of the beta-glucan product was determined by gel permeation chromatography in combination with multi-angle laser light scattering and refractive index detector (white tech company of sababara, ca, usa). Sodium nitrate (0.1M) and sodium azide (0.02% w/v) were used as mobile phases at a rate of 0.4 mL/min. The temperature of the column (Shodex Ohpak SB-805 HQ) and refractive index detector was controlled at 60 ℃. The measurement data were analyzed by the Astra 6.0 software package.
Claims (8)
1. The green and efficient highland barley beta-glucan extracting process includes the following steps:
(1) Grinding highland barley into powder; grinding semen Avenae Nudae to obtain powder;
(2) Ethanol extraction: extracting highland barley powder with 80% ethanol, and oven drying the precipitate;
(3) Leaching with hot water: extracting with distilled water, centrifuging, and mixing the supernatants;
(4) Enzyme removal: sequentially adding high temperature resistant alpha-amylase, saccharifying enzyme and pancreatin into the supernatant for enzymolysis, inactivating enzyme, centrifuging, and concentrating the supernatant;
(5) Crude extraction: and after alcohol precipitation, centrifugally washing and freeze-drying to obtain the beta-glucan extract.
2. The method for green and efficient extraction of highland barley beta-glucan according to claim 1, wherein the highland barley powder and 80% ethanol in the step (2) are mixed according to the following formula 1:10w/v, the temperature of reflux extraction is 80 ℃ and the time is 2h.
3. The method for extracting highland barley beta-glucan with high efficiency according to claim 1, wherein in the step (3), distilled water at 52 ℃ is used for extracting 2h, and the extraction is repeated three times, wherein the feed-liquid ratio is 1:10w/v.
4. The method for efficiently extracting highland barley beta-glucan according to claim 1, wherein the high temperature resistant alpha-amylase in the step (4) is added in an amount of 10U/g, the pH value is 6.5, and enzymolysis is carried out for 2 hours at 95 ℃.
5. The method for extracting highland barley beta-glucan with high efficiency according to claim 1, wherein the adding amount of saccharifying enzyme in the step (4) is 200U/g, the pH value is 4.5, and enzymolysis is carried out at 60 ℃ for 30min.
6. The method for efficiently extracting highland barley beta-glucan according to claim 1, wherein the adding amount of pancreatin in the step (4) is 200U/g, the pH value is 4.5, and enzymolysis is carried out for 1h at 50 ℃.
7. The method for efficiently extracting highland barley beta-glucan according to claim 1, wherein the centrifugation conditions in the step (3) are 5000rpm for 10min, and the centrifugation conditions in the step (4) and the step (5) are 5000rpm for 15min.
8. The method for green and efficient extraction of highland barley beta-glucan according to claim 1, wherein the concentration of ethanol in the final solution required for the ethanol precipitation in the step (8) is 80%.
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Cited By (1)
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN117859917A (en) * | 2023-12-29 | 2024-04-12 | 北京联合大学 | Preparation with memory improving capability and preparation method thereof |
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