WO2023205915A1 - Preparation process of sargassum fusiforme oligosaccharide and use of sargassum fusiforme oligosaccharide in crop planting - Google Patents

Preparation process of sargassum fusiforme oligosaccharide and use of sargassum fusiforme oligosaccharide in crop planting Download PDF

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Publication number
WO2023205915A1
WO2023205915A1 PCT/CN2022/088651 CN2022088651W WO2023205915A1 WO 2023205915 A1 WO2023205915 A1 WO 2023205915A1 CN 2022088651 W CN2022088651 W CN 2022088651W WO 2023205915 A1 WO2023205915 A1 WO 2023205915A1
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Prior art keywords
hijiki
oligosaccharide
crops
application according
tobacco
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PCT/CN2022/088651
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French (fr)
Chinese (zh)
Inventor
李义强
陈芊如
荆常亮
邹平
袁源
马斯琦
李振
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中国农业科学院烟草研究所
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Priority to PCT/CN2022/088651 priority Critical patent/WO2023205915A1/en
Publication of WO2023205915A1 publication Critical patent/WO2023205915A1/en

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    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
    • A01G22/00—Cultivation of specific crops or plants not otherwise provided for
    • A01G22/45—Tobacco
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
    • A01G7/00—Botany in general
    • A01G7/06—Treatment of growing trees or plants, e.g. for preventing decay of wood, for tingeing flowers or wood, for prolonging the life of plants
    • C—CHEMISTRY; METALLURGY
    • C05—FERTILISERS; MANUFACTURE THEREOF
    • C05G—MIXTURES OF FERTILISERS COVERED INDIVIDUALLY BY DIFFERENT SUBCLASSES OF CLASS C05; MIXTURES OF ONE OR MORE FERTILISERS WITH MATERIALS NOT HAVING A SPECIFIC FERTILISING ACTIVITY, e.g. PESTICIDES, SOIL-CONDITIONERS, WETTING AGENTS; FERTILISERS CHARACTERISED BY THEIR FORM
    • C05G3/00—Mixtures of one or more fertilisers with additives not having a specially fertilising activity

Definitions

  • the invention belongs to the field of crop planting and relates to the preparation technology of hijiki oligosaccharide and its application in crop planting.
  • the purpose of the present invention is to provide the application of hijiki oligosaccharide in crop planting.
  • the hijiki oligosaccharide can promote the growth of crops and improve the quality of crops.
  • the invention provides the application of hijiki oligosaccharide as fertilizer in crop planting.
  • the crop cultivation is potted cultivation, and the application includes using hijiki oligosaccharide solution to fertilize potted crops;
  • the mass concentration of hijiki oligosaccharide in the hijiki oligosaccharide solution is 0.01g/L to 0.05g/L.
  • the fertilization includes root irrigation and foliar spraying.
  • the fertilization includes: starting from the 3rd day of transplanting the crops, performing the root irrigation three times continuously, with the time interval between two adjacent root irrigations being 5 days, and the amount of application for each root irrigation being 50 mL/plant;
  • the foliar spraying was carried out 4 times continuously.
  • the time interval between two adjacent foliar sprayings was 5d, and the application amount of each foliar spraying was ⁇ 6.67mL/plant.
  • the crop planting is field planting of crops, and the application includes using Hijiki oligosaccharide solution to fertilize field crops;
  • the mass concentration of the hijiki oligosaccharide in the hijiki oligosaccharide solution is 0.1g/L to 0.5g/L.
  • the fertilization includes root irrigation.
  • the fertilization includes: performing the root irrigation twice or three times continuously when the crops enter the vigorous period.
  • the time interval between two adjacent root irrigations is 7 to 14 days, and the amount of application for each root irrigation is 50 mL. /strain.
  • the preparation method of hijiki oligosaccharide includes the following steps:
  • the hijiki polysaccharide is subjected to a second enzymatic hydrolysis using alginic acid lyase to obtain hijiki oligosaccharide.
  • the ratio of the mass sum of the cellulase, pectinase and papain to the dry weight of the hijiki is 5 to 7:100.
  • the mass ratio of cellulase, pectinase and papain is 2-3:2-3:1.
  • the temperature of the first enzymatic hydrolysis is 50-60°C, the time is 60-180 min, and the pH value is 6-8.
  • the mass ratio of the alginate lyase to the hijiki polysaccharide is 2-3:100.
  • the temperature of the second enzymatic hydrolysis is 50-60°C, the time is 60-180 min, and the pH value is 6-8.
  • the crops include tobacco.
  • the tobacco is planted in large and small rows, with the row spacing of the large rows being 1.4m and the row spacing of the small rows being 1.2m; the tobacco plant spacing is 50cm.
  • the hijiki oligosaccharide of the present invention has the advantages of promoting growth, improving quality, being green and non-toxic to crops, can improve the antioxidant capacity of crops, enhance the agronomic traits and nutritional characteristics of crops, and can especially improve the plant height, The number of leaves per plant, leaf length, leaf width, leaf area, node spacing and fresh weight, and during the growth process of crops, the contents of total chlorophyll, total sugar, total phenols and total flavonoids in crop leaves were significantly increased.
  • the embodiments of the present invention take tobacco as an example to illustrate the growth-promoting and quality-improving effects of hijiki oligosaccharides on crops.
  • the yield of tobacco and The output value increased by 13.24% to 14.56% and 18.21% to 20.09% respectively; the total nicotine content and chloride ion content of tobacco treated with hijiki oligosaccharide, total sugar, reducing sugar, two sugar ratios and sugar-alkali ratio contents all increased.
  • the starch content also increased significantly.
  • the hijiki oligosaccharide of the present invention can significantly improve the quality of tobacco leaves, reduce the irritation of tobacco, increase the aroma and taste of tobacco leaves, and improve the economic and social benefits of tobacco leaves.
  • the invention provides the application of hijiki oligosaccharide as fertilizer in crop planting.
  • the hijiki oligosaccharide of the present invention is preferably used as a growth stimulant type fertilizer.
  • the hijiki oligosaccharide can be purchased or prepared by oneself.
  • the preparation method of the hijiki oligosaccharide preferably includes the following steps:
  • the hijiki polysaccharide is subjected to a second enzymatic hydrolysis using alginic acid lyase to obtain hijiki oligosaccharide.
  • the sieve size used is preferably 60 mesh, so as to obtain hijiki powder with a particle size of ⁇ 60 mesh.
  • the present invention has no requirement on the source of the hijiki, and it can be purchased directly. For example, hijiki products can be purchased directly.
  • the present invention uses cellulase, pectinase and papain to perform the first enzymatic hydrolysis of the hijiki powder to obtain hijiki polysaccharide.
  • the ratio of the total mass of the cellulase, pectinase and papain to the mass of the hijiki powder is preferably 5 to 7:100, and further preferably 6:100.
  • the mass ratio of cellulase, pectinase and papain is preferably 2-3:2-3:1, and more preferably 3:3:1.
  • the first enzymatic hydrolysis time is preferably 60-180 min, more preferably 65-115 min or 120-175 min, more preferably 75-105 min or 125-160 min, most preferably 85-95 min or 130-130 min. 145min;
  • the temperature of the first enzymatic hydrolysis is preferably 50-60°C, more preferably 52-58°C, more preferably 55°C;
  • the pH value of the first enzymatic hydrolysis is preferably 6-8, further preferably 7 .
  • the first enzymatic hydrolysis of hijiki using cellulase, pectinase and papain in the present invention preferably includes: mixing cellulase, pectinase and papain to obtain a composite enzyme;
  • the composite enzyme is mixed with hijiki and water to perform the first enzymatic hydrolysis.
  • the enzyme activity of the cellulase is preferably 20,000 U/g; the enzyme activity of the pectinase is preferably 30,000 U/g; and the enzyme activity of the papain is preferably 80 10,000 U/g.
  • the present invention does not have strict requirements on the source of the cellulase, pectinase and papain, and they can be purchased.
  • the cellulase, pectinase and papain were purchased from Nanning Pangbo Bioengineering Co., Ltd.
  • the mass volume ratio of the hijiki powder and water is preferably 1g:40mL.
  • the present invention preferably performs post-processing on the obtained first enzymatic hydrolysis liquid to obtain hijiki polysaccharide.
  • the first enzymatic hydrolyzate is centrifuged, and the supernatant is an extract containing hijiki polysaccharide.
  • the centrifugal speed of the present invention is preferably 3000-8000r/min, more preferably 4000-7000r/min, more preferably 5000-6000r/min; the centrifugation time is preferably 3-5min, further preferably 4min.
  • the present invention utilizes cellulase, pectinase and papain to enzymatically hydrolyze hijiki.
  • the embodiments of the present invention control the dosage of cellulase, pectinase and papain, the time, temperature and pH value of enzymatic hydrolysis. , capable of efficiently enzymatically hydrolyzing hijiki to obtain 76.4-96.74mg/g hijiki polysaccharide.
  • the present invention uses alginic acid lyase to perform a second enzymatic hydrolysis on the hijiki polysaccharide or the extract containing the hijiki polysaccharide to obtain hijiki oligosaccharide.
  • the mass ratio of the alginic acid lyase to the hijiki polysaccharide is preferably 2 to 3:100; in one embodiment, the mass ratio of the alginic acid lyase to the hijiki polysaccharide is The added amount of rapeseed polysaccharide can be consistent with the added amount of cellulase relative to hijiki in the first enzymatic hydrolysis process.
  • the second enzymatic hydrolysis time is preferably 60-180 min, more preferably 65-115 min or 120-175 min, more preferably 75-105 min or 125-160 min, most preferably 85-95 min or 130-130 min. 145min;
  • the temperature of the second enzymatic hydrolysis is preferably 50-60°C, more preferably 52-58°C, more preferably 55°C;
  • the pH value of the second enzymatic hydrolysis is preferably 6-8, further preferably 7 .
  • the present invention does not have strict requirements on the source of the alginate lyase, and any alginate lyase produced by any manufacturer on the market can be used.
  • Embodiments of the present invention utilize alginic acid lyase to enzymatically hydrolyze hijiki polysaccharide, and by controlling the dosage of alginic acid lyase, the time, temperature and pH value of enzymatic hydrolysis, the hijiki polysaccharide can be degraded to obtain Oligosaccharide polymers within 20 monosaccharide residues, that is, hijiki oligosaccharides with a molecular weight of ⁇ 3600 Da, are easy for plants to absorb.
  • the present invention preferably performs post-processing on the obtained second enzymatic hydrolysis solution to obtain Hijiki oligosaccharide.
  • the second enzymatic hydrolyzate is preferably freeze-dried to obtain the hijiki oligosaccharide.
  • the present invention does not have strict requirements on the freeze-drying method, and conventional freeze-drying methods can be used.
  • the hijiki oligosaccharide can be applied to the cultivation of any variety of crops, and is especially suitable for the cultivation of tobacco.
  • the planting described in the present invention can be potted planting or field planting.
  • the hijiki oligosaccharide of the present invention is used for potted crops, it is preferred that the hijiki oligosaccharide is formulated into a hijiki oligosaccharide aqueous solution, and the hijiki oligosaccharide solution is used to fertilize the potted crops.
  • the plant seedlings grown in pots according to the present invention preferably include seedlings that are cultivated in a seedbed and then transplanted into a planting pot.
  • one crop plant is preferably grown in each cultivation pot.
  • the diameter of the planting pot of the present invention is preferably 9 cm.
  • the present invention does not have strict requirements on the method of cultivating the seedbed, and it can be carried out in a routine manner.
  • the mass concentration of the hijiki oligosaccharide in the hijiki oligosaccharide solution is 0.01g/L to 0.05g/L, and more preferably 0.02g/L to 0.04g/L.
  • Fertilization according to the present invention preferably includes root irrigation and foliar spraying.
  • the time for two consecutive root irrigations is preferably 5 days; each time the root irrigation is performed, the application amount of the hijiki oligosaccharide solution is preferably 50 mL/plant.
  • the present invention preferably starts foliar spraying 5 days after the last root irrigation treatment, and the foliar spraying is preferably carried out four times in a row.
  • the time interval between two adjacent foliar sprayings in the present invention is preferably 5 days; each time the foliar spraying is performed, the application amount of the hijiki oligosaccharide solution is preferably ⁇ 6.67mL/plant.
  • the hijiki oligosaccharide of the present invention is preferably formulated into a hijiki oligosaccharide solution, and the hijiki oligosaccharide solution is used to fertilize field crops.
  • tobacco is used as an example to illustrate.
  • the row spacing of tobacco planting is preferably divided into large and small rows.
  • the row spacing of large rows is preferably 1.4m, and the row spacing of small rows is preferably 1.4m.
  • the distance between plants is 1.2m, and the distance between plants is preferably 50cm.
  • the mass concentration of the hijiki oligosaccharide in the hijiki oligosaccharide aqueous solution is 0.1g/L to 0.5g/L, and more preferably 0.2 to 0.4g. /L.
  • Fertilization according to the present invention preferably includes root irrigation.
  • the time interval between two consecutive root irrigations in the present invention is preferably 7 to 14 days, and further preferably 8 to 10 days; each time the root irrigation is performed, the application amount of the hijiki oligosaccharide solution is preferably 50 mL/ strain.
  • pure nitrogen and nitrogen, phosphorus and potassium compound fertilizers are applied according to the local conventional tobacco leaf production standards.
  • the hijiki oligosaccharide obtained by the above preparation method of the present invention has a small molecular weight, is easy to be absorbed by plants, especially has growth-promoting and quality-improving effects on crops, is green and non-toxic, can be used for crop planting, and can improve the antioxidant capacity of crops. , improve the quality of crops, have a significant positive effect on promoting early growth and vigorous growth of crops, and can improve the economic and social benefits of crops.
  • tobacco is used as a representative crop to verify the growth-promoting effect of the hijiki oligosaccharide on crops of the present invention, but it cannot be limited to the fact that the hijiki oligosaccharide of the present invention only affects tobacco. It has a promoting effect on growth and constitution.
  • Hijiki oligosaccharide can increase the plant height of tobacco, the number of leaves per plant, leaf length, leaf width, leaf area, pitch and fresh weight, and during the growth process of tobacco, the total chlorophyll, The contents of total sugar, total phenols and total flavonoids were significantly increased.
  • the yield and output value of tobacco increased by 13.24% to 14.56% and 18.21% to 20.09% respectively; the total nicotine content and chloride ion content of tobacco after treatment with hijiki oligosaccharide, total sugar, reducing sugar, two sugar ratio and sugar base The specific content increased, and the starch content also increased significantly.
  • Pretreatment Dry the purchased hijiki products, grind them, pass them through a 60-mesh sieve, and the part below the sieve will be dry hijiki powder.
  • step (2) the enzymatic hydrolysis solution is centrifuged for 5 minutes at a rotation speed of 4000 r/min, and the supernatant is collected to obtain the hijiki polysaccharide extract.
  • step (3) Determine the polysaccharide content in the hijiki polysaccharide extract obtained in step (3) (the results are shown in Table 2), add 3% alginic acid lyase in the polysaccharide mass of the hijiki polysaccharide extract, and adjust the pH value to 7 , perform a second enzymatic hydrolysis at 50°C for 180 minutes to obtain a hijiki oligosaccharide degradation solution.
  • Hijiki oligosaccharide is prepared according to the method of Example 1, wherein the addition amounts of cellulase, pectinase and papain in Examples 2 to 4 are the same as those in Example 1; enzymatic hydrolysis time of steps (2) of Examples 4 to 6 Same as Example 1; the added amounts of cellulase, pectinase and papain in Examples 6 to 8 are the same; the added amounts of cellulase, pectinase and papain in Examples 9 to 10 are the same as in Example 5. The details are listed in Table 1 below.
  • the content of hijiki polysaccharide in the hijiki polysaccharide extract obtained in step (3) of Examples 1 to 10 was measured.
  • the measurement results are as follows in Table 2.
  • step (1) Use a seedling tray to cultivate 1,000 small golden tobacco seeds.
  • the tobacco seedlings grow to 4 leaves and 1 heart, select strong and consistent seedlings and transplant them into flowerpots with a diameter of 9cm.
  • Each flowerpot Transplant a tobacco seedling.
  • the tobacco seedlings are treated according to step (2). Each treatment contains 15 pots of tobacco seedlings.
  • foliar spraying was started to fertilize the tobacco seedlings of different treatments.
  • spraying try to ensure that the spray dosage of each tobacco seedling is roughly the same, and spray until the leaves are dripping with water.
  • the spraying dose of each treatment shall not exceed 100mL, that is, the spraying dose of each tobacco plant shall not exceed 6.67mL.
  • the fertilization interval is 5 days, and fertilization is sprayed 4 times in total. All treatments were watered regularly during the rest of the time.
  • hijiki oligosaccharide has a significant promoting effect on the plant height, number of leaves per plant, leaf length, leaf width and fresh weight of potted tobacco, and can promote the growth of tobacco.
  • Step (2) Weeks after the last foliar fertilization, cut the leaves from the same part of each treatment into pieces, put them into a mortar, add an appropriate amount of liquid nitrogen, grind them, and use ethanol with a volume concentration of 95% as the solvent. Cut an appropriate amount of tobacco leaves into pieces, put them into a mortar, add an appropriate amount of liquid nitrogen, and grind them into pieces. After weighing 0.1g, add 5 mL of 95% ethanol and vortex to form a homogenate. Centrifuge the homogenate at 8000r/min for 8 minutes, take 500 ⁇ L of the supernatant, add it to a new 2mL tube, then add 1.5mL of 95% ethanol, and mix.
  • the absorbance values were measured at 440nm, 649nm and 665nm respectively, and the chlorophyll content and soluble sugar content of each treatment were determined; the leaves of each treatment were freeze-dried and ground with a grinder, and the total phenols and total flavonoids of each treatment were determined. content, the measurement results are shown in Table 4.
  • chlorophyll calculation formula is as follows:
  • Chlorophyll a concentration (mg/L) 12.71 ⁇ A 665 -2.59 ⁇ A 649
  • Chlorophyll b concentration (mg/L) 22.88 ⁇ A 649 -4.67 ⁇ A 665
  • Hijiki oligosaccharides can improve the antioxidant capacity of tobacco, and have a significant role in promoting the early growth and growth of tobacco, promoting growth and quality.
  • the altitude is 36 meters, with one acre as the standard, large and small rows are planted, the large row spacing is 1.4m, the small row spacing is 1.2m, the plant spacing is 50cm, the number of tobacco plants is 1026 plants/acre, and pure nitrogen is applied at 92.4kg/hm 2 , nitrogen, phosphorus and potassium.
  • Other cultivation measures are implemented in accordance with Shandongzhou flue-cured tobacco production technical specifications.
  • the Hijiki oligosaccharide obtained in Example 1 is mixed with water at a concentration of 0.1g/L and 0.5g/L to obtain Hijiki oligosaccharide.
  • the tobacco seedlings were treated with root irrigation and fertilization using vegetable oligosaccharide aqueous solution.
  • three treatment plots were set up: 1 Clear water control (CK); 2 Hijiki oligosaccharide aqueous solution 0.1g/L; 3 Hijiki oligosaccharide aqueous solution 0.5 g/L, a total of 50 tobacco plants in each treatment plot, three experiments were set up in each treatment plot, and all treatment plots were randomly arranged.
  • 100 mL was applied to the roots of each flue-cured tobacco plant, and the fertilization interval was 10 days, with a total of 3 fertilizations.
  • the ratio of two sugars is reducing sugar/total sugar.
  • the 0.1g/L hijiki oligosaccharide treatment group resulted in a decrease in the total nicotine content in tobacco leaves, an increase in the reducing sugar content, an increase in the two sugar ratios, and a decrease in chloride ion content; the 0.5g/L hijiki oligosaccharide treatment group
  • the total nicotine content decreased, the total sugar and reducing sugar content increased, the chloride ion content decreased, and the two-sugar ratio and the sugar-alkali ratio increased.
  • the starch content of both treatment groups increased significantly, indicating that both can significantly improve tobacco leaf quality.
  • Hijiki oligosaccharides have a significant improvement effect on tobacco, can increase the aroma and taste of tobacco leaves, reduce the irritation of tobacco leaves, and thus significantly increase the quality and output value of tobacco leaves.
  • the hijiki oligosaccharide of the present invention can promote the growth of tobacco, significantly improve the quality of tobacco leaves, reduce the irritation of tobacco leaves, increase the aroma and taste of tobacco leaves, and improve the economic and social benefits of tobacco leaves.

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Abstract

Disclosed are a preparation process of sargassum fusiforme oligosaccharide and the use of the sargassum fusiforme oligosaccharide in crop planting. The sargassum fusiforme oligosaccharide has the advantages for crops, such as promoting growth, improving quality, being green and non-toxic, can be used as a fertilizer for crop field planting and potted plant planting, improves the antioxidant capacity of crops, enhances the agronomic traits and nutritional characteristics of crops, improves the quality of crops, increases the yield of crops, and increases economic benefits and social benefits.

Description

羊栖菜寡糖制备工艺及在农作物种植中的应用Preparation technology of hijiki oligosaccharide and its application in crop planting 技术领域Technical field
本发明属于农作物种植领域，涉及羊栖菜寡糖制备技术及在农作物种植中的应用。The invention belongs to the field of crop planting and relates to the preparation technology of hijiki oligosaccharide and its application in crop planting.
背景技术Background technique
长期以来农作物种植过程中依赖化肥的农业生产模式，给农业生态环境、作物生长造成巨大压力，农产品品质与对病虫害的抗性亟待提升。在种植农作物时，施肥和施用植物刺激素是非常关键的环节。目前，市场以及研究领域通常是在农作物种植过程中添加相关植物刺激素，以提高农作物的产量和品质，如中国专利CN108191583A公开了在烟草肥料中加入氯化锌、硝酸钾等的方案，其在烟草生长中施加中微量元素，提高了烟草的产量，同时改善了烟草的质量和土质结构。但是中微量元素以离子形式存在，一方面会有相互之间的拮抗作用，影响吸收；另一方面，直接进入烟田会与土壤中阴离子形成难溶性盐，严重影响中微量元素的施用效果及其利用率，并没有达到中微量元素对于烟草生长应有的作用。The long-term agricultural production model that relies on chemical fertilizers during crop planting has put huge pressure on the agricultural ecological environment and crop growth. The quality of agricultural products and resistance to pests and diseases need to be improved urgently. When growing crops, fertilization and application of plant stimulants are very critical links. At present, the market and research fields usually add related plant stimulants during the crop planting process to improve the yield and quality of crops. For example, Chinese patent CN108191583A discloses a solution for adding zinc chloride, potassium nitrate, etc. to tobacco fertilizer. The application of medium and trace elements during tobacco growth increases tobacco yield, while improving tobacco quality and soil structure. However, the medium and trace elements exist in the form of ions. On the one hand, there will be mutual antagonism and affect the absorption; on the other hand, if they enter the tobacco field directly, they will form insoluble salts with the anions in the soil, seriously affecting the application effect and effectiveness of the medium and trace elements. Its utilization rate has not reached the expected role of medium and trace elements in tobacco growth.
因此，如何通过施用植物源刺激素等特色肥料促进农作物的生长，提高农作物的质量，仍是农作物种植领域亟待解决的问题。Therefore, how to promote the growth of crops and improve the quality of crops through the application of special fertilizers such as phytostimulants is still an urgent problem to be solved in the field of crop planting.
发明内容Contents of the invention
本发明的目的在于提供羊栖菜寡糖在农作物种植中的应用，羊栖菜寡糖能够促进农作物的生长，提高农作物质量。The purpose of the present invention is to provide the application of hijiki oligosaccharide in crop planting. The hijiki oligosaccharide can promote the growth of crops and improve the quality of crops.
本发明提供了羊栖菜寡糖在农作物种植中作为肥料的应用。The invention provides the application of hijiki oligosaccharide as fertilizer in crop planting.
优选的，所述农作物种植为盆栽种植，所述应用包括，利用羊栖菜寡糖溶液对盆栽农作物施肥；Preferably, the crop cultivation is potted cultivation, and the application includes using hijiki oligosaccharide solution to fertilize potted crops;
所述羊栖菜寡糖溶液中羊栖菜寡糖的质量浓度为0.01g/L～0.05g/L。The mass concentration of hijiki oligosaccharide in the hijiki oligosaccharide solution is 0.01g/L to 0.05g/L.
优选的，所述施肥包括灌根和叶面喷施。Preferably, the fertilization includes root irrigation and foliar spraying.
优选的，所述施肥包括：在移栽农作物第3d开始，连续进行3次所述灌根，相邻两次灌根的时间间隔为5d，每次灌根的施用量为50mL/株；Preferably, the fertilization includes: starting from the 3rd day of transplanting the crops, performing the root irrigation three times continuously, with the time interval between two adjacent root irrigations being 5 days, and the amount of application for each root irrigation being 50 mL/plant;
在最后一次灌根处理5d后开始，连续进行4次所述叶面喷施，相邻两次叶面喷施的时间间隔为5d，每次叶面喷施的施用量≤6.67mL/株。Starting 5 days after the last root irrigation treatment, the foliar spraying was carried out 4 times continuously. The time interval between two adjacent foliar sprayings was 5d, and the application amount of each foliar spraying was ≤6.67mL/plant.
优选的，所述农作物种植为农作物大田种植，所述应用包括，采用羊栖菜 寡糖溶液对大田农作物施肥；Preferably, the crop planting is field planting of crops, and the application includes using Hijiki oligosaccharide solution to fertilize field crops;
所述羊栖菜寡糖溶液中羊栖菜寡糖的质量浓度为0.1g/L～0.5g/L。The mass concentration of the hijiki oligosaccharide in the hijiki oligosaccharide solution is 0.1g/L to 0.5g/L.
优选的，所述施肥包括灌根。Preferably, the fertilization includes root irrigation.
优选的，所述施肥包括：在农作物进入旺长期开始，连续进行2次或3次所述灌根，相邻两次灌根的时间间隔为7～14d，每次灌根的施用量为50mL/株。Preferably, the fertilization includes: performing the root irrigation twice or three times continuously when the crops enter the vigorous period. The time interval between two adjacent root irrigations is 7 to 14 days, and the amount of application for each root irrigation is 50 mL. /strain.
优选的，所述羊栖菜寡糖的制备方法，包括如下步骤：Preferably, the preparation method of hijiki oligosaccharide includes the following steps:
利用纤维素酶、果胶酶和木瓜蛋白酶对羊栖菜进行第一酶解，得羊栖菜多糖；Use cellulase, pectinase and papain to perform the first enzymatic hydrolysis of hijiki to obtain hijiki polysaccharide;
利用褐藻酸裂解酶对所述羊栖菜多糖进行第二酶解，得羊栖菜寡糖。The hijiki polysaccharide is subjected to a second enzymatic hydrolysis using alginic acid lyase to obtain hijiki oligosaccharide.
优选的，所述纤维素酶、果胶酶和木瓜蛋白酶的质量和与所述羊栖菜干重的比例为5～7:100。Preferably, the ratio of the mass sum of the cellulase, pectinase and papain to the dry weight of the hijiki is 5 to 7:100.
优选的，所述纤维素酶、果胶酶和木瓜蛋白酶的质量比为2～3:2～3:1。Preferably, the mass ratio of cellulase, pectinase and papain is 2-3:2-3:1.
优选的，所述第一酶解的温度为50～60℃，时间为60～180min，pH值为6～8。Preferably, the temperature of the first enzymatic hydrolysis is 50-60°C, the time is 60-180 min, and the pH value is 6-8.
优选的，所述褐藻酸裂解酶与所述羊栖菜多糖的质量比为2～3:100。Preferably, the mass ratio of the alginate lyase to the hijiki polysaccharide is 2-3:100.
优选的，所述第二酶解的温度为50～60℃，时间为60～180min，pH值为6～8。Preferably, the temperature of the second enzymatic hydrolysis is 50-60°C, the time is 60-180 min, and the pH value is 6-8.
优选的，所述农作物包括烟草。Preferably, the crops include tobacco.
优选的，所述烟草分大小行种植，大行的行距为1.4m，小行的行距为1.2m；所述烟草的株距为50cm。Preferably, the tobacco is planted in large and small rows, with the row spacing of the large rows being 1.4m and the row spacing of the small rows being 1.2m; the tobacco plant spacing is 50cm.
本发明所述羊栖菜寡糖对农作物具有促生、提质、绿色无毒等优势，能够提高农作物抗氧化能力，增强农作物的农艺性状和营养特性，尤其能够提高烟草等农作物的株高、单株叶片数、叶长、叶宽、叶面积、节距和鲜重，并且在农作物生长过程中，农作物叶片总叶绿素、总糖、总酚和总黄酮的含量均有明显提高。本发明实施例以烟草为例说明羊栖菜寡糖对农作物的促生提质作用，结果表明，羊栖菜寡糖对促进烟株早发旺长等有明显正向作用，烟草的产量和产值分别提高13.24％～14.56％和18.21％～20.09％；经羊栖菜寡糖处理后的烟草总烟碱含量和氯离子含量，总糖、还原糖、两糖比和糖碱比含量均上升，淀粉含量也有显著提高。本发明羊栖菜寡糖可以显著提升烟叶品质，降低烟草的刺激性，增加烟叶香气吃味，提高烟叶经济效益以及社会效益。The hijiki oligosaccharide of the present invention has the advantages of promoting growth, improving quality, being green and non-toxic to crops, can improve the antioxidant capacity of crops, enhance the agronomic traits and nutritional characteristics of crops, and can especially improve the plant height, The number of leaves per plant, leaf length, leaf width, leaf area, node spacing and fresh weight, and during the growth process of crops, the contents of total chlorophyll, total sugar, total phenols and total flavonoids in crop leaves were significantly increased. The embodiments of the present invention take tobacco as an example to illustrate the growth-promoting and quality-improving effects of hijiki oligosaccharides on crops. The results show that hijiki oligosaccharides have a significant positive effect on promoting early growth and vigorous growth of tobacco plants. The yield of tobacco and The output value increased by 13.24% to 14.56% and 18.21% to 20.09% respectively; the total nicotine content and chloride ion content of tobacco treated with hijiki oligosaccharide, total sugar, reducing sugar, two sugar ratios and sugar-alkali ratio contents all increased. , the starch content also increased significantly. The hijiki oligosaccharide of the present invention can significantly improve the quality of tobacco leaves, reduce the irritation of tobacco, increase the aroma and taste of tobacco leaves, and improve the economic and social benefits of tobacco leaves.
具体实施方式Detailed ways
本发明提供了羊栖菜寡糖在农作物种植中作为肥料的应用。本发明所述羊栖菜寡糖优选作为生长刺激素类特色肥料使用。The invention provides the application of hijiki oligosaccharide as fertilizer in crop planting. The hijiki oligosaccharide of the present invention is preferably used as a growth stimulant type fertilizer.
在本发明中，所述羊栖菜寡糖可购买获得，或者自行制备。在本发明的实施例中，所述羊栖菜寡糖的制备方法优选包括如下步骤：In the present invention, the hijiki oligosaccharide can be purchased or prepared by oneself. In the embodiment of the present invention, the preparation method of the hijiki oligosaccharide preferably includes the following steps:
利用纤维素酶、果胶酶和木瓜蛋白酶对羊栖菜进行第一酶解，得羊栖菜多糖；Use cellulase, pectinase and papain to perform the first enzymatic hydrolysis of hijiki to obtain hijiki polysaccharide;
利用褐藻酸裂解酶对所述羊栖菜多糖进行第二酶解，得羊栖菜寡糖。The hijiki polysaccharide is subjected to a second enzymatic hydrolysis using alginic acid lyase to obtain hijiki oligosaccharide.
本发明优选进行所述第一酶解前，对羊栖菜进行烘干和粉碎过筛处理，得羊栖菜粉末。本发明进行所述过筛处理时，使用的筛孔大小优选为60目，获得粒径≤60目的羊栖菜粉末。本发明对所述羊栖菜的来源没有要求，可直接购买获得，例如可以直接购买羊栖菜制品。In the present invention, it is preferred to dry, crush and sieve the hijiki before performing the first enzymatic hydrolysis to obtain hijiki powder. When the present invention performs the sieving process, the sieve size used is preferably 60 mesh, so as to obtain hijiki powder with a particle size of ≤60 mesh. The present invention has no requirement on the source of the hijiki, and it can be purchased directly. For example, hijiki products can be purchased directly.
得羊栖菜粉末后，本发明利用纤维素酶、果胶酶和木瓜蛋白酶对羊栖菜粉末进行第一酶解，得羊栖菜多糖。在本发明中，所述纤维素酶、果胶酶和木瓜蛋白酶的总质量和与所述羊栖菜粉末质量的比优选为5～7:100，进一步优选为6:100。在本发明中，所述纤维素酶、果胶酶和木瓜蛋白酶的质量比优选为2～3:2～3:1，进一步优选为3:3:1。After obtaining the hijiki powder, the present invention uses cellulase, pectinase and papain to perform the first enzymatic hydrolysis of the hijiki powder to obtain hijiki polysaccharide. In the present invention, the ratio of the total mass of the cellulase, pectinase and papain to the mass of the hijiki powder is preferably 5 to 7:100, and further preferably 6:100. In the present invention, the mass ratio of cellulase, pectinase and papain is preferably 2-3:2-3:1, and more preferably 3:3:1.
在本发明中，所述第一酶解的时间优选为60～180min，进一步优选为65～115min或120～175min，更优选为75～105min或125～160min，最优选为85～95min或130～145min；所述第一酶解的温度优选为50～60℃，进一步优选为52～58℃，更优选为55℃；所述第一酶解的pH值优选为6～8，进一步优选为7。In the present invention, the first enzymatic hydrolysis time is preferably 60-180 min, more preferably 65-115 min or 120-175 min, more preferably 75-105 min or 125-160 min, most preferably 85-95 min or 130-130 min. 145min; the temperature of the first enzymatic hydrolysis is preferably 50-60°C, more preferably 52-58°C, more preferably 55°C; the pH value of the first enzymatic hydrolysis is preferably 6-8, further preferably 7 .
本发明所述利用纤维素酶、果胶酶和木瓜蛋白酶对羊栖菜进行第一酶解优选包括：将纤维素酶、果胶酶和木瓜蛋白酶混合，得复合酶；The first enzymatic hydrolysis of hijiki using cellulase, pectinase and papain in the present invention preferably includes: mixing cellulase, pectinase and papain to obtain a composite enzyme;
将所述复合酶与羊栖菜和水混合进行所述第一酶解。The composite enzyme is mixed with hijiki and water to perform the first enzymatic hydrolysis.
本发明所述复合酶中，所述纤维素酶的酶活力优选为2万U/g；所述果胶酶的酶活力优选为3万U/g；所述木瓜蛋白酶的酶活力优选为80万U/g。本发明对所述纤维素酶、果胶酶和木瓜蛋白酶的来源没有严格要求，购买获得即可。本发明具体实施过程中，所述纤维素酶、果胶酶和木瓜蛋白酶均购买自南宁庞博生物工程有限公司。In the composite enzyme of the present invention, the enzyme activity of the cellulase is preferably 20,000 U/g; the enzyme activity of the pectinase is preferably 30,000 U/g; and the enzyme activity of the papain is preferably 80 10,000 U/g. The present invention does not have strict requirements on the source of the cellulase, pectinase and papain, and they can be purchased. During the specific implementation of the present invention, the cellulase, pectinase and papain were purchased from Nanning Pangbo Bioengineering Co., Ltd.
本发明优选将羊栖菜粉末与水混合，对得到的溶液体系进行第一酶解。在本发明中，所述羊栖菜粉末和水的质量体积比优选为1g：40mL。In the present invention, it is preferred to mix hijiki powder and water, and perform the first enzymatic hydrolysis on the resulting solution system. In the present invention, the mass volume ratio of the hijiki powder and water is preferably 1g:40mL.
所述第一酶解后，本发明优选将得到的第一酶解液进行后处理，得到羊栖菜多糖。本发明优选对所述第一酶解液进行离心处理，上清液为含有羊栖菜多糖的提取液。本发明所述离心的转速优选为3000～8000r/min，进一步优选为 4000～7000r/min，更优选为5000～6000r/min；所述离心的时间优选为3～5min，进一步优选为4min。After the first enzymatic hydrolysis, the present invention preferably performs post-processing on the obtained first enzymatic hydrolysis liquid to obtain hijiki polysaccharide. In the present invention, it is preferred that the first enzymatic hydrolyzate is centrifuged, and the supernatant is an extract containing hijiki polysaccharide. The centrifugal speed of the present invention is preferably 3000-8000r/min, more preferably 4000-7000r/min, more preferably 5000-6000r/min; the centrifugation time is preferably 3-5min, further preferably 4min.
本发明利用纤维素酶、果胶酶和木瓜蛋白酶酶解羊栖菜，本发明的实施例控制纤维素酶、果胶酶和木瓜蛋白酶的用量，酶解的时间、温度和酶解的pH值，能够使羊栖菜高效酶解，得到76.4～96.74mg/g羊栖菜多糖。The present invention utilizes cellulase, pectinase and papain to enzymatically hydrolyze hijiki. The embodiments of the present invention control the dosage of cellulase, pectinase and papain, the time, temperature and pH value of enzymatic hydrolysis. , capable of efficiently enzymatically hydrolyzing hijiki to obtain 76.4-96.74mg/g hijiki polysaccharide.
得羊栖菜多糖后，本发明利用褐藻酸裂解酶对所述羊栖菜多糖或所述含有羊栖菜多糖的提取液进行第二酶解，得羊栖菜寡糖。本发明在具体实施过程中，所述褐藻酸裂解酶的质量与所述羊栖菜多糖的质量比优选为2～3:100；在一个实施例中，褐藻酸裂解酶相对于所述羊栖菜多糖的添加量可以与第一酶解过程中所述纤维素酶相对于羊栖菜的添加量一致。After obtaining the hijiki polysaccharide, the present invention uses alginic acid lyase to perform a second enzymatic hydrolysis on the hijiki polysaccharide or the extract containing the hijiki polysaccharide to obtain hijiki oligosaccharide. During the specific implementation of the present invention, the mass ratio of the alginic acid lyase to the hijiki polysaccharide is preferably 2 to 3:100; in one embodiment, the mass ratio of the alginic acid lyase to the hijiki polysaccharide is The added amount of rapeseed polysaccharide can be consistent with the added amount of cellulase relative to hijiki in the first enzymatic hydrolysis process.
在本发明中，所述第二酶解的时间优选为60～180min，进一步优选为65～115min或120～175min，更优选为75～105min或125～160min，最优选为85～95min或130～145min；所述第二酶解的温度优选为50～60℃，进一步优选为52～58℃，更优选为55℃；所述第二酶解的pH值优选为6～8，进一步优选为7。In the present invention, the second enzymatic hydrolysis time is preferably 60-180 min, more preferably 65-115 min or 120-175 min, more preferably 75-105 min or 125-160 min, most preferably 85-95 min or 130-130 min. 145min; the temperature of the second enzymatic hydrolysis is preferably 50-60°C, more preferably 52-58°C, more preferably 55°C; the pH value of the second enzymatic hydrolysis is preferably 6-8, further preferably 7 .
本发明对所述褐藻酸裂解酶的来源没有严格要求，市面上任何厂家生产的褐藻酸裂解酶均可。本发明的实施例利用褐藻酸裂解酶酶解羊栖菜多糖，并且通过控制纤褐藻酸裂解酶的用量，酶解的时间、温度和酶解的pH值，能够将羊栖菜多糖降解，得20个单糖残基以内的低聚糖类聚合物，即分子量≤3600Da的羊栖菜寡糖，易于植物吸收。The present invention does not have strict requirements on the source of the alginate lyase, and any alginate lyase produced by any manufacturer on the market can be used. Embodiments of the present invention utilize alginic acid lyase to enzymatically hydrolyze hijiki polysaccharide, and by controlling the dosage of alginic acid lyase, the time, temperature and pH value of enzymatic hydrolysis, the hijiki polysaccharide can be degraded to obtain Oligosaccharide polymers within 20 monosaccharide residues, that is, hijiki oligosaccharides with a molecular weight of ≤3600 Da, are easy for plants to absorb.
所述第二酶解后，本发明优选将得到的第二酶解液进行后处理，得到羊栖菜寡糖。本发明优选将所述第二酶解液进行冻干处理，得所述羊栖菜寡糖。本发明对所述冻干处理的方式没有严格要求，采用常规的冻干方式即可。After the second enzymatic hydrolysis, the present invention preferably performs post-processing on the obtained second enzymatic hydrolysis solution to obtain Hijiki oligosaccharide. In the present invention, the second enzymatic hydrolyzate is preferably freeze-dried to obtain the hijiki oligosaccharide. The present invention does not have strict requirements on the freeze-drying method, and conventional freeze-drying methods can be used.
在本发明中，所述羊栖菜寡糖能够应用于任何品种的农作物的种植，尤其适用于烟草的种植。In the present invention, the hijiki oligosaccharide can be applied to the cultivation of any variety of crops, and is especially suitable for the cultivation of tobacco.
本发明所述种植可以为盆栽种植或大田种植。当本发明所述羊栖菜寡糖用于农作物盆栽种植时，优选将所述羊栖菜寡糖配制成羊栖菜寡糖水溶液，利用所述羊栖菜寡糖溶液对盆栽农作物施肥。本发明所述盆栽种植的植物苗优选包括经苗床培育后移栽至栽种盆中的秧苗。本发明优选经苗床培育后长到4叶1心时，进行移栽，实现农作物的盆栽种植。本发明优选在每个栽培盆中种植一株农作物。本发明所述栽种盆的直径优选为9cm。本发明对所述苗床培育的方式没有严格要求，常规进行即可。The planting described in the present invention can be potted planting or field planting. When the hijiki oligosaccharide of the present invention is used for potted crops, it is preferred that the hijiki oligosaccharide is formulated into a hijiki oligosaccharide aqueous solution, and the hijiki oligosaccharide solution is used to fertilize the potted crops. The plant seedlings grown in pots according to the present invention preferably include seedlings that are cultivated in a seedbed and then transplanted into a planting pot. In the present invention, it is preferable to transplant crops when they grow to 4 leaves and 1 heart after being cultivated in a seedbed to realize potted planting of crops. In the present invention, one crop plant is preferably grown in each cultivation pot. The diameter of the planting pot of the present invention is preferably 9 cm. The present invention does not have strict requirements on the method of cultivating the seedbed, and it can be carried out in a routine manner.
在本发明中，所述盆栽种植时，所述羊栖菜寡糖溶液中羊栖菜寡糖的质量浓度为0.01g/L～0.05g/L，进一步优选为0.02g/L～0.04g/L。本发明所述施肥优选包括灌根和叶面喷施。本发明优选在移栽农作物第3d开始，连续进行3次所述灌根。本发明相邻两次所述灌根的时间优选为5d；每次进行所述灌根时，所述羊栖菜寡糖溶液的施用量优选为50mL/株。In the present invention, when the potted plant is grown, the mass concentration of the hijiki oligosaccharide in the hijiki oligosaccharide solution is 0.01g/L to 0.05g/L, and more preferably 0.02g/L to 0.04g/L. L. Fertilization according to the present invention preferably includes root irrigation and foliar spraying. In the present invention, it is preferable to perform the root irrigation three times continuously starting from the 3rd day after transplanting the crops. In the present invention, the time for two consecutive root irrigations is preferably 5 days; each time the root irrigation is performed, the application amount of the hijiki oligosaccharide solution is preferably 50 mL/plant.
灌根结束后，本发明优选自最后一次灌根处理5d后开始叶面喷施，所述叶面喷施优选为连续进行4次叶面喷施。本发明相邻两次所述叶面喷施的时间间隔优选为5d；每次进行所述叶面喷施时，所述羊栖菜寡糖溶液的施用量优选≤6.67mL/株。After the root irrigation is completed, the present invention preferably starts foliar spraying 5 days after the last root irrigation treatment, and the foliar spraying is preferably carried out four times in a row. The time interval between two adjacent foliar sprayings in the present invention is preferably 5 days; each time the foliar spraying is performed, the application amount of the hijiki oligosaccharide solution is preferably ≤6.67mL/plant.
在本发明中，进行所述盆栽种植时，除本发明限定的灌根和叶面喷施的施肥方式外，盆栽种植过程中涉及到的其他处理，如土壤的选择，浇水量等均按照当地盆栽常规农作物生产标准配合进行。In the present invention, when planting potted plants, in addition to the fertilization methods of root irrigation and foliar spraying as defined in the invention, other treatments involved in the potted planting process, such as soil selection, watering amount, etc., are all in accordance with Local potted conventional crop production standards are carried out.
当本发明所述羊栖菜寡糖用于农作物大田种植时，优选将所述羊栖菜寡糖配制成羊栖菜寡糖溶液，利用所述羊栖菜寡糖溶液对大田农作物施肥。本发明在具体实施过程中，以烟草例进行说明，具体的，在烟草大田种植中，所述烟草栽种的株行距优选分大小行种植，大行的行距优选为1.4m，小行的行距优选为1.2m，株距优选为50cm。在选择其他农作物时，以符合相应农作物品种的种植模式选择最佳株行距规格即可。When the hijiki oligosaccharide of the present invention is used for field planting of crops, the hijiki oligosaccharide is preferably formulated into a hijiki oligosaccharide solution, and the hijiki oligosaccharide solution is used to fertilize field crops. In the specific implementation process of the present invention, tobacco is used as an example to illustrate. Specifically, in tobacco field planting, the row spacing of tobacco planting is preferably divided into large and small rows. The row spacing of large rows is preferably 1.4m, and the row spacing of small rows is preferably 1.4m. The distance between plants is 1.2m, and the distance between plants is preferably 50cm. When selecting other crops, just select the best plant-row spacing specifications based on the planting pattern that matches the corresponding crop variety.
在本发明中，当所述农作物种植为大田种植时，所述羊栖菜寡糖水溶液中羊栖菜寡糖的质量浓度为0.1g/L～0.5g/L，进一步优选为0.2～0.4g/L。本发明所述施肥优选包括灌根。本发明优选自农作物进入旺盛生长阶段开始，连续进行2次或3次所述灌根。本发明相邻两次所述灌根的时间间隔优选为7～14d，进一步优选为8～10d；每次进行所述灌根时，所述羊栖菜寡糖溶液的施用量优选为50mL/株。In the present invention, when the crops are grown in field cultivation, the mass concentration of the hijiki oligosaccharide in the hijiki oligosaccharide aqueous solution is 0.1g/L to 0.5g/L, and more preferably 0.2 to 0.4g. /L. Fertilization according to the present invention preferably includes root irrigation. In the present invention, it is preferable to perform the root irrigation twice or three times continuously from the time when the crops enter the vigorous growth stage. The time interval between two consecutive root irrigations in the present invention is preferably 7 to 14 days, and further preferably 8 to 10 days; each time the root irrigation is performed, the application amount of the hijiki oligosaccharide solution is preferably 50 mL/ strain.
在本发明中，进行所述大田种植时，按照当地大田常规烟叶生产标准配合施加纯氮和氮磷钾复合肥。本发明所述氮磷钾复合肥中氮磷钾的质量比优选为N：P 2O 5：K 2O＝1.00：0.72：3.83。 In the present invention, during field planting, pure nitrogen and nitrogen, phosphorus and potassium compound fertilizers are applied according to the local conventional tobacco leaf production standards. The mass ratio of nitrogen, phosphorus and potassium in the nitrogen, phosphorus and potassium compound fertilizer of the present invention is preferably N: P2O5 : K2O =1.00:0.72:3.83.
本发明利用上述制备方法得到的羊栖菜寡糖分子量小，易于被植物吸收，尤其对农作物具有促生、提质的作用，并且绿色无毒，能够用于农作物种植，能够提高农作物抗氧化能力，提升农作物品质，对促进农作物早发旺长等有明显正向作用，可以提高农作物经济效益以及社会效益。The hijiki oligosaccharide obtained by the above preparation method of the present invention has a small molecular weight, is easy to be absorbed by plants, especially has growth-promoting and quality-improving effects on crops, is green and non-toxic, can be used for crop planting, and can improve the antioxidant capacity of crops. , improve the quality of crops, have a significant positive effect on promoting early growth and vigorous growth of crops, and can improve the economic and social benefits of crops.
本发明在具体实施过程中，以烟草为农作物代表，验证本发明所述羊栖菜寡糖对农作物的促生体质作用，但是不能将其限定为本发明所述羊栖菜寡糖只对烟草的促生和体质有促进作用。本发明实施例结果表明，羊栖菜寡糖能够提高烟草的株高，单株叶片数，叶长，叶宽，叶面积，节距和鲜重，并且烟草在烟草生长过程中，总叶绿素、总糖、总酚和总黄酮的含量均有明显提高。烟草的产量和产值分别提高13.24％～14.56％和18.21％～20.09％；经羊栖菜寡糖处理后的烟草总烟碱含量和氯离子含量，总糖、还原糖、两糖比和糖碱比含量均上升，淀粉含量也有显著提高。During the specific implementation of the present invention, tobacco is used as a representative crop to verify the growth-promoting effect of the hijiki oligosaccharide on crops of the present invention, but it cannot be limited to the fact that the hijiki oligosaccharide of the present invention only affects tobacco. It has a promoting effect on growth and constitution. The results of the embodiments of the present invention show that Hijiki oligosaccharide can increase the plant height of tobacco, the number of leaves per plant, leaf length, leaf width, leaf area, pitch and fresh weight, and during the growth process of tobacco, the total chlorophyll, The contents of total sugar, total phenols and total flavonoids were significantly increased. The yield and output value of tobacco increased by 13.24% to 14.56% and 18.21% to 20.09% respectively; the total nicotine content and chloride ion content of tobacco after treatment with hijiki oligosaccharide, total sugar, reducing sugar, two sugar ratio and sugar base The specific content increased, and the starch content also increased significantly.
为了进一步说明本发明，下面结合实施例对本发明提供的技术方案进行详细地描述，但不能将它们理解为对本发明保护范围的限定。In order to further illustrate the present invention, the technical solutions provided by the present invention are described in detail below in conjunction with the examples, but they should not be understood as limiting the protection scope of the present invention.
实施例1Example 1
羊栖菜寡糖的制备步骤如下：The preparation steps of hijiki oligosaccharide are as follows:
(1)预处理：将采购的羊栖菜制品烘干后磨碎，过60目筛，筛下部分为栖菜干粉。(1) Pretreatment: Dry the purchased hijiki products, grind them, pass them through a 60-mesh sieve, and the part below the sieve will be dry hijiki powder.
(2)酶解：取1g羊栖菜干粉加入50mL的离心管中，按羊栖菜干粉与水的质量体积比1g：40mL将水与羊栖菜干粉混合，加入羊栖菜干粉质量3％的纤维素酶、3％的果胶酶和1％的木瓜蛋白酶，调节pH值为7，在50℃第一酶解180min。(2) Enzymatic hydrolysis: Add 1g of hijiki dry powder into a 50mL centrifuge tube, mix the water and hijiki dry powder according to the mass volume ratio of hijiki dry powder and water: 1g:40mL, and add 3% of the mass of hijiki dry powder Cellulase, 3% pectinase and 1% papain, adjust the pH value to 7, and perform the first enzymatic hydrolysis at 50°C for 180 minutes.
(3)离心：步骤(2)酶解溶液在转速为4000r/min条件下离心5min，收集上清液，得羊栖菜多糖提取液。(3) Centrifugation: In step (2), the enzymatic hydrolysis solution is centrifuged for 5 minutes at a rotation speed of 4000 r/min, and the supernatant is collected to obtain the hijiki polysaccharide extract.
(4)测定步骤(3)所得羊栖菜多糖提取液中多糖的含量(结果如表2所示)，加入羊栖菜多糖提取液中多糖质量3％褐藻酸裂解酶，调节pH值为7，在50℃第二酶解180min，得羊栖菜寡糖降解液。(4) Determine the polysaccharide content in the hijiki polysaccharide extract obtained in step (3) (the results are shown in Table 2), add 3% alginic acid lyase in the polysaccharide mass of the hijiki polysaccharide extract, and adjust the pH value to 7 , perform a second enzymatic hydrolysis at 50°C for 180 minutes to obtain a hijiki oligosaccharide degradation solution.
(5)冻干：取步骤(4)所得羊栖菜寡糖降解液，冻干得羊栖菜寡糖。(5) Freeze-drying: Take the hijiki oligosaccharide degradation solution obtained in step (4) and freeze-dry it to obtain hijiki oligosaccharide.
实施例2～10Examples 2 to 10
按照实施例1的方法制备羊栖菜寡糖，其中实施例2～4纤维素酶，果胶酶和木瓜蛋白酶的添加量与实施例1相同；实施例4～6步骤(2)酶解时间与实施例1相同；实施例6～8纤维素酶，果胶酶和木瓜蛋白酶的添加量相同；实施例9～10纤维素酶，果胶酶和木瓜蛋白酶的添加量与实施例5相同，具体列于下表1中。Hijiki oligosaccharide is prepared according to the method of Example 1, wherein the addition amounts of cellulase, pectinase and papain in Examples 2 to 4 are the same as those in Example 1; enzymatic hydrolysis time of steps (2) of Examples 4 to 6 Same as Example 1; the added amounts of cellulase, pectinase and papain in Examples 6 to 8 are the same; the added amounts of cellulase, pectinase and papain in Examples 9 to 10 are the same as in Example 5. The details are listed in Table 1 below.
表1实施例1～10不同参数的设定Table 1 Settings of different parameters in Embodiments 1 to 10
Figure PCTCN2022088651-appb-000001
Figure PCTCN2022088651-appb-000001
测定实施例1～10步骤(3)所得羊栖菜多糖提取液中羊栖菜多糖的含量，测定结果如下表2。The content of hijiki polysaccharide in the hijiki polysaccharide extract obtained in step (3) of Examples 1 to 10 was measured. The measurement results are as follows in Table 2.
表2实施例1～10羊栖菜多糖提取液中羊栖菜多糖的质量Table 2 The quality of hijiki polysaccharide in the hijiki polysaccharide extract solution of Examples 1 to 10
Figure PCTCN2022088651-appb-000002
Figure PCTCN2022088651-appb-000002
由表2可以看出，本发明通过严格控制纤维素酶、果胶酶和木瓜蛋白酶的用量，酶解的时间、温度和酶解的pH值，能够使羊栖菜高效酶解，得到76.4～96.74mg/g羊栖菜多糖，提高酶解效果。It can be seen from Table 2 that by strictly controlling the dosage of cellulase, pectinase and papain, the time, temperature and pH value of enzymatic hydrolysis, the present invention can efficiently enzymatically hydrolyze hijiki to obtain 76.4~ 96.74mg/g Hijiki polysaccharide improves enzymatic hydrolysis effect.
应用例1盆栽种植Application Example 1 Potted Planting
(1)用育苗盘培育1000棵小黄金烟草种子。在出苗后1周，将烟苗移栽到72孔育苗盘中，在烟苗长到4叶1心时，选取健壮、长势一致的幼苗移栽到直径为9cm花盆中，每个花盆移栽一株烟草幼苗，烟苗移栽3d后按照步骤(2)处理方式进行处理，每个处理包含15盆烟苗。(1) Use a seedling tray to cultivate 1,000 small golden tobacco seeds. One week after emergence, transplant the tobacco seedlings into a 72-hole seedling tray. When the tobacco seedlings grow to 4 leaves and 1 heart, select strong and consistent seedlings and transplant them into flowerpots with a diameter of 9cm. Each flowerpot Transplant a tobacco seedling. After 3 days of transplanting, the tobacco seedlings are treated according to step (2). Each treatment contains 15 pots of tobacco seedlings.
(2)以0.01g/L、0.05g/L的浓度，将实施例1获得的羊栖菜寡糖添加到水中，得到羊栖菜寡糖水溶液，对烟苗进行施肥处理，具体的，共设3个处理：共设3个处理：①清水对照(CK)；②羊栖菜寡糖水溶液0.01g/L；③羊栖菜寡糖水溶液0.05g/L。采用灌根法对不同处理的烟苗进行施肥处理，每盆剂量为50mL，每隔5d一次，共3次。在最后一次灌根处理5d后，开始采取叶面喷施对不同处理的烟苗进行施肥处理，喷施时尽量保证每株烟苗的喷施剂量大致相同，喷施至叶片滴水的程度即可，每个处理的喷施剂量不超过100mL，即每株烟草的喷施剂量不超过6.67mL。施肥间隔时间为5d，共喷施施肥4次。其余时间所有处理按期浇水。(2) Add the hijiki oligosaccharide obtained in Example 1 to water at a concentration of 0.01g/L and 0.05g/L to obtain a hijiki oligosaccharide aqueous solution, and fertilize the tobacco seedlings. Specifically, Set up 3 treatments: Set up 3 treatments in total: ① Clear water control (CK); ② Hijiki oligosaccharide aqueous solution 0.01g/L; ③ Hijiki oligosaccharide aqueous solution 0.05g/L. The tobacco seedlings of different treatments were fertilized using the root irrigation method. The dose per pot was 50 mL, once every 5 days, a total of 3 times. 5 days after the last root irrigation treatment, foliar spraying was started to fertilize the tobacco seedlings of different treatments. When spraying, try to ensure that the spray dosage of each tobacco seedling is roughly the same, and spray until the leaves are dripping with water. , the spraying dose of each treatment shall not exceed 100mL, that is, the spraying dose of each tobacco plant shall not exceed 6.67mL. The fertilization interval is 5 days, and fertilization is sprayed 4 times in total. All treatments were watered regularly during the rest of the time.
最后一次叶面施肥结束后一周，取样、测定烟叶农艺性状，测定结果见表3。One week after the last foliar fertilization, the agronomic properties of the tobacco leaves were sampled and measured. The measurement results are shown in Table 3.
表3不同处理下烟草叶部农艺性状Table 3 Agronomic traits of tobacco leaves under different treatments
Figure PCTCN2022088651-appb-000003
Figure PCTCN2022088651-appb-000003
由表3可以看出，羊栖菜寡糖对盆栽烟草的株高，单株叶片数，叶长，叶宽和鲜重都有明显的促进作用，能够促进烟草生长。It can be seen from Table 3 that hijiki oligosaccharide has a significant promoting effect on the plant height, number of leaves per plant, leaf length, leaf width and fresh weight of potted tobacco, and can promote the growth of tobacco.
步骤(2)最后一次叶面施肥结束周后，取每个处理中相同部位的叶片剪碎，放入研钵中，加入适量液氮，研磨碎，以体积浓度95％的乙醇为溶剂，取适量烟草叶片剪碎，放入研钵中，加入适量液氮，研磨碎。称取0.1g后，加入5mL95％乙醇，涡旋成匀浆。将匀浆液在8000r/min下离心8min，取上清液500μL，加入到一个新的2mL管里，再加入1.5mL 95％乙醇，混匀。分别在440nm，649nm和665nm下测定吸光度值，测定各个处理的叶绿素含量及可溶性糖含量；将每个处理的叶片进行冷冻干燥后，用研磨机磨碎，测定各个处理的总酚、总黄酮的含量，测定结果见表4。Step (2) Weeks after the last foliar fertilization, cut the leaves from the same part of each treatment into pieces, put them into a mortar, add an appropriate amount of liquid nitrogen, grind them, and use ethanol with a volume concentration of 95% as the solvent. Cut an appropriate amount of tobacco leaves into pieces, put them into a mortar, add an appropriate amount of liquid nitrogen, and grind them into pieces. After weighing 0.1g, add 5 mL of 95% ethanol and vortex to form a homogenate. Centrifuge the homogenate at 8000r/min for 8 minutes, take 500μL of the supernatant, add it to a new 2mL tube, then add 1.5mL of 95% ethanol, and mix. The absorbance values were measured at 440nm, 649nm and 665nm respectively, and the chlorophyll content and soluble sugar content of each treatment were determined; the leaves of each treatment were freeze-dried and ground with a grinder, and the total phenols and total flavonoids of each treatment were determined. content, the measurement results are shown in Table 4.
其中，叶绿素计算公式为如下：Among them, the chlorophyll calculation formula is as follows:
叶绿素a浓度(mg/L)＝12.71×A 665-2.59×A 649 Chlorophyll a concentration (mg/L)=12.71×A 665 -2.59×A 649
叶绿素b浓度(mg/L)＝22.88×A 649-4.67×A 665 Chlorophyll b concentration (mg/L)=22.88×A 649 -4.67×A 665
叶绿素总含量(mg/L)＝C a+C b Total chlorophyll content (mg/L)=C a +C b
表4不同处理下烟草烟草营养生长特性Table 4 Tobacco tobacco nutritional growth characteristics under different treatments
Figure PCTCN2022088651-appb-000004
Figure PCTCN2022088651-appb-000004
由表4可以看出，羊栖菜寡糖处理组盆栽烟叶绿素含量、可溶性糖含量都比对照有显著增加，且总酚、总黄酮也显著增加。羊栖菜寡糖能够提高烟草的抗氧化能力，对烟草早发快长、促生提质有显著的促进作用。It can be seen from Table 4 that the chlorophyll content and soluble sugar content of potted tobacco in the hijiki oligosaccharide treatment group were significantly increased compared with the control, and the total phenols and total flavonoids were also significantly increased. Hijiki oligosaccharides can improve the antioxidant capacity of tobacco, and have a significant role in promoting the early growth and growth of tobacco, promoting growth and quality.
应用例2大田种植Application example 2 field planting
试验于山东省青岛市黄岛区大村烟站进行，土壤类型为壤土。海拔36米，以一亩为标准，大小行种植，大行行距为1.4m，小行行距1.2m，株距50cm，植烟株数1026株/亩，施纯氮92.4kg/hm 2，氮磷钾施肥比为m(N)：m(P 2O 5)：m(K 2O)＝1.00：0.72：3.83，其他栽培措施均按山东省烤烟生产技术规范执行。 The experiment was conducted at the Dacun Tobacco Station in Huangdao District, Qingdao City, Shandong Province, and the soil type was loam. The altitude is 36 meters, with one acre as the standard, large and small rows are planted, the large row spacing is 1.4m, the small row spacing is 1.2m, the plant spacing is 50cm, the number of tobacco plants is 1026 plants/acre, and pure nitrogen is applied at 92.4kg/hm 2 , nitrogen, phosphorus and potassium. The fertilization ratio is m(N):m(P 2 O 5 ): m(K 2 O) = 1.00: 0.72: 3.83. Other cultivation measures are implemented in accordance with Shandong Province flue-cured tobacco production technical specifications.
在烟苗长到团棵期后，将实施例1获得的羊栖菜寡糖以0.1g/L、0.5g/L的浓度实施例1获得的羊栖菜寡糖与水混合，得羊栖菜寡糖水溶液对烟苗进行灌根施肥处理，具体的，共设3个处理小区：①清水对照(CK)；②羊栖菜寡糖水溶液0.1g/L；③羊栖菜寡糖水溶液0.5g/L，每个处理小区共计50株烟草，每个处理小区设置3次实验，所有处理小区随机排列。每个处理小区每株烤烟灌根施入100mL，施肥间隔时间为10d，共施肥3次。After the tobacco seedlings grow to the group stage, the Hijiki oligosaccharide obtained in Example 1 is mixed with water at a concentration of 0.1g/L and 0.5g/L to obtain Hijiki oligosaccharide. The tobacco seedlings were treated with root irrigation and fertilization using vegetable oligosaccharide aqueous solution. Specifically, three treatment plots were set up: ① Clear water control (CK); ② Hijiki oligosaccharide aqueous solution 0.1g/L; ③ Hijiki oligosaccharide aqueous solution 0.5 g/L, a total of 50 tobacco plants in each treatment plot, three experiments were set up in each treatment plot, and all treatment plots were randomly arranged. In each treatment plot, 100 mL was applied to the roots of each flue-cured tobacco plant, and the fertilization interval was 10 days, with a total of 3 fertilizations.
于大田成熟期(移栽后60d左右)，每个处理小区随机测量6株烟株的主要农艺性状，包括株高、有效叶数、茎围、最大叶长叶宽和最大叶面积。采收后，测定C3F烟叶的经济性状和化学品质分析指标，测定结果如下表5～7。At the maturity stage of the field (about 60 days after transplanting), the main agronomic traits of 6 tobacco plants in each treatment plot were randomly measured, including plant height, number of effective leaves, stem circumference, maximum leaf length and width, and maximum leaf area. After harvesting, the economic properties and chemical quality analysis indicators of C3F tobacco leaves were measured. The measurement results are as follows in Tables 5 to 7.
表5不同处理下大田种植烤烟叶部农艺性状Table 5 Agronomic properties of flue-cured tobacco leaves grown in field under different treatments
Figure PCTCN2022088651-appb-000005
Figure PCTCN2022088651-appb-000005
Figure PCTCN2022088651-appb-000006
Figure PCTCN2022088651-appb-000006
根据表5可以看出，羊栖菜寡糖0.1g/L处理组烤烟的株高显著高于对照组，羊栖菜寡糖0.5g/L处理组的株高、有效叶数、茎围都显著高于对照组，羊栖菜寡糖对大田烟草有显著的促生作用。According to Table 5, it can be seen that the plant height of flue-cured tobacco in the hijiki oligosaccharide 0.1g/L treatment group was significantly higher than that in the control group. The plant height, effective leaf number, and stem circumference of the hijiki oligosaccharide 0.5g/L treatment group were all the same. Significantly higher than the control group, Hijiki oligosaccharide has a significant growth-promoting effect on field tobacco.
表6不同处理下烤烟叶部化学成分含量及协调性的影响Table 6 Effects of chemical composition content and coordination of flue-cured tobacco leaves under different treatments
Figure PCTCN2022088651-appb-000007
Figure PCTCN2022088651-appb-000007
注：两糖比为还原糖/总糖。Note: The ratio of two sugars is reducing sugar/total sugar.
根据表6可以看出，羊栖菜寡糖0.1g/L处理组和0.5g/L处理组施加羊栖菜寡糖降解液后，烤烟叶的化学成分发生了较为显著的改变，其中，羊栖菜寡糖0.1g/L处理组的导致烟草叶片中的总烟碱含量降低，还原糖含量上升，两糖比上升，此外氯离子含量下降；羊栖菜寡糖0.5g/L处理组的总烟碱含量下降，总糖和还原糖含量均上升，氯离子含量下降，提高了两糖比、糖碱比。同时，两个处理组的淀粉含量均显著上升，表明二者均可以显著提升烟叶品质。羊栖菜寡糖对烟草有显著的提质作用，能够增加烟叶香气吃味，降低烟叶的刺激性，进而显著增加烟叶品质和产值。According to Table 6, it can be seen that the chemical composition of flue-cured tobacco leaves has changed significantly after the application of Hijiki oligosaccharide degradation liquid in the 0.1g/L and 0.5g/L treatment groups of Hijiki oligosaccharide. Among them, the chemical composition of Hijiki oligosaccharide has changed significantly. The 0.1g/L hijiki oligosaccharide treatment group resulted in a decrease in the total nicotine content in tobacco leaves, an increase in the reducing sugar content, an increase in the two sugar ratios, and a decrease in chloride ion content; the 0.5g/L hijiki oligosaccharide treatment group The total nicotine content decreased, the total sugar and reducing sugar content increased, the chloride ion content decreased, and the two-sugar ratio and the sugar-alkali ratio increased. At the same time, the starch content of both treatment groups increased significantly, indicating that both can significantly improve tobacco leaf quality. Hijiki oligosaccharides have a significant improvement effect on tobacco, can increase the aroma and taste of tobacco leaves, reduce the irritation of tobacco leaves, and thus significantly increase the quality and output value of tobacco leaves.
表7不同处理下烟草产量Table 7 Tobacco yield under different treatments
Figure PCTCN2022088651-appb-000008
Figure PCTCN2022088651-appb-000008
根据表7可以看出，施加羊栖菜寡糖后，烟草的经济性状普遍增加，羊栖菜寡糖0.1g/L处理组的烤烟均价上升，产量产值显著增加，且上等烟比例也升 高；羊栖菜寡糖0.5g/L处理组的烤烟的均价、产量产值以及上等烟比例均显著高于对照组，表明羊栖菜寡糖对烟草有显著的提质和增产作用，能够大大增加烟叶产量产值。According to Table 7, it can be seen that after the application of hijiki oligosaccharide, the economic traits of tobacco generally increased. The average price of flue-cured tobacco in the hijiki oligosaccharide 0.1g/L treatment group increased, the output value increased significantly, and the proportion of high-quality tobacco also increased. increased; the average price, yield value and proportion of high-quality tobacco in the hijiki oligosaccharide 0.5g/L treatment group were significantly higher than those in the control group, indicating that hijiki oligosaccharide has a significant effect on improving tobacco quality and increasing yield. , can greatly increase the output value of tobacco leaf production.
由以上述内容可知，本发明羊栖菜寡糖可以促进烟草生长，显著提升烟叶品质，降低烟叶的刺激性，增加烟叶香气吃味，提高烟叶经济效益以及社会效益。It can be seen from the above that the hijiki oligosaccharide of the present invention can promote the growth of tobacco, significantly improve the quality of tobacco leaves, reduce the irritation of tobacco leaves, increase the aroma and taste of tobacco leaves, and improve the economic and social benefits of tobacco leaves.
尽管上述实施例对本发明做出了详尽的描述，但它仅仅是本发明一部分实施例，而不是全部实施例，人们还可以根据本实施例在不经创造性前提下获得其他实施例，这些实施例都属于本发明保护范围。Although the above embodiments describe the present invention in detail, they are only some of the embodiments of the present invention, not all of them. People can also obtain other embodiments based on this embodiment without any inventive step. These embodiments All belong to the protection scope of the present invention.

Claims (15)

  1. 羊栖菜寡糖在农作物种植中作为肥料的应用。Application of hijiki oligosaccharide as fertilizer in crop cultivation.
  2. 根据权利要求1所述的应用，其特征在于，所述农作物种植为盆栽种植，所述应用包括，利用羊栖菜寡糖溶液对盆栽农作物施肥；The application according to claim 1, characterized in that the crop planting is potted planting, and the application includes using hijiki oligosaccharide solution to fertilize the potted crops;
    所述羊栖菜寡糖溶液中羊栖菜寡糖的质量浓度为0.01g/L～0.05g/L。The mass concentration of hijiki oligosaccharide in the hijiki oligosaccharide solution is 0.01g/L to 0.05g/L.
  3. 根据权利要求2所述的应用，其特征在于，所述施肥包括灌根和叶面喷施。The application according to claim 2, characterized in that the fertilization includes root irrigation and foliar spraying.
  4. 根据权利要求3所述的应用，其特征在于，所述施肥包括：在移栽农作物第3d开始，连续进行3次所述灌根，相邻两次灌根的时间间隔为5d，每次灌根的施用量为50mL/株；The application according to claim 3, characterized in that the fertilization includes: starting from the third day of transplanting the crops, performing the root irrigation three times continuously, and the time interval between two adjacent root irrigations is 5 days, and each irrigation The application amount for roots is 50mL/plant;
    在最后一次灌根处理5d后开始，连续进行4次所述叶面喷施，相邻两次叶面喷施的时间间隔为5d，每次叶面喷施的施用量≤6.67mL/株。Starting 5 days after the last root irrigation treatment, the foliar spraying was carried out 4 times continuously. The time interval between two adjacent foliar sprayings was 5d, and the application amount of each foliar spraying was ≤6.67mL/plant.
  5. 根据权利要求1所述的应用，其特征在于，所述农作物种植为农作物大田种植，所述应用包括，采用羊栖菜寡糖溶液对大田农作物施肥；The application according to claim 1, characterized in that the crop planting is field planting of crops, and the application includes using Hijiki oligosaccharide solution to fertilize field crops;
    所述羊栖菜寡糖溶液中羊栖菜寡糖的质量浓度为0.1g/L～0.5g/L。The mass concentration of the hijiki oligosaccharide in the hijiki oligosaccharide solution is 0.1g/L to 0.5g/L.
  6. 根据权利要求5所述的应用，其特征在于，所述施肥包括灌根。The application according to claim 5, characterized in that said fertilizing includes root irrigation.
  7. 根据权利要求6所述的应用，其特征在于，所述施肥包括：在农作物进入旺长期开始，连续进行2次或3次所述灌根，相邻两次灌根的时间间隔为7～14d，每次灌根的施用量为50mL/株。The application according to claim 6, characterized in that the fertilization includes: when the crops enter a prosperous period, the root irrigation is performed 2 or 3 times continuously, and the time interval between two adjacent root irrigations is 7 to 14 days. , the application amount for each root irrigation is 50mL/plant.
  8. 根据权利要求1所述的应用，其特征在于，所述羊栖菜寡糖的制备方法，包括如下步骤：The application according to claim 1, characterized in that the preparation method of the hijiki oligosaccharide includes the following steps:
    利用纤维素酶、果胶酶和木瓜蛋白酶对羊栖菜进行第一酶解，得羊栖菜多糖；Use cellulase, pectinase and papain to perform the first enzymatic hydrolysis of hijiki to obtain hijiki polysaccharide;
    利用褐藻酸裂解酶对所述羊栖菜多糖进行第二酶解，得羊栖菜寡糖。The hijiki polysaccharide is subjected to a second enzymatic hydrolysis using alginic acid lyase to obtain hijiki oligosaccharide.
  9. 根据权利要求8所述的应用，其特征在于，所述纤维素酶、果胶酶和木瓜蛋白酶的质量和与所述羊栖菜干重的比例为5～7:100。The application according to claim 8, characterized in that the ratio of the mass sum of the cellulase, pectinase and papain to the dry weight of the hijiki is 5 to 7:100.
  10. 根据权利要求9所述的应用，其特征在于，所述纤维素酶、果胶酶和木瓜蛋白酶的质量比为2～3:2～3:1。The application according to claim 9, characterized in that the mass ratio of cellulase, pectinase and papain is 2-3:2-3:1.
  11. 根据权利要求8所述的应用，其特征在于，所述第一酶解的温度为50～60℃，时间为60～180min，pH值为6～8。The application according to claim 8, characterized in that the temperature of the first enzymatic hydrolysis is 50-60°C, the time is 60-180 min, and the pH value is 6-8.
  12. 根据权利要求8所述的应用，其特征在于，所述褐藻酸裂解酶与所述羊 栖菜多糖的质量比为2～3:100。The application according to claim 8, characterized in that the mass ratio of the alginic acid lyase and the hijiki polysaccharide is 2 to 3:100.
  13. 根据权利要求8所述的应用，其特征在于，所述第二酶解的温度为50～60℃，时间为60～180min，pH值为6～8。The application according to claim 8, characterized in that the temperature of the second enzymatic hydrolysis is 50-60°C, the time is 60-180 min, and the pH value is 6-8.
  14. 根据权利要求1～13任一项所述的应用，其特征在于，所述农作物包括烟草。The application according to any one of claims 1 to 13, characterized in that the crops include tobacco.
  15. 根据权利要求14所述的应用，其特征在于，所述烟草分大小行种植，大行的行距为1.4m，小行的行距为1.2m；所述烟草的株距为50cm。The application according to claim 14, characterized in that the tobacco is planted in large rows, the row spacing of the large rows is 1.4m, and the row spacing of the small rows is 1.2m; the tobacco plant spacing is 50cm.
PCT/CN2022/088651 2022-04-24 2022-04-24 Preparation process of sargassum fusiforme oligosaccharide and use of sargassum fusiforme oligosaccharide in crop planting WO2023205915A1 (en)

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