WO2018161361A1 - 一种采用生物工程技术培植的鱼饵保鲜剂的制备方法 - Google Patents

一种采用生物工程技术培植的鱼饵保鲜剂的制备方法 Download PDF

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WO2018161361A1
WO2018161361A1 PCT/CN2017/076405 CN2017076405W WO2018161361A1 WO 2018161361 A1 WO2018161361 A1 WO 2018161361A1 CN 2017076405 W CN2017076405 W CN 2017076405W WO 2018161361 A1 WO2018161361 A1 WO 2018161361A1
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preservative
bacteria
total number
preparation
water
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PCT/CN2017/076405
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English (en)
French (fr)
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耿胜利
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耿胜利
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Priority to DE112017002924.4T priority Critical patent/DE112017002924T5/de
Publication of WO2018161361A1 publication Critical patent/WO2018161361A1/zh

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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L17/00—Food-from-the-sea products; Fish products; Fish meal; Fish-egg substitutes; Preparation or treatment thereof
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L3/00—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs
    • A23L3/34—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals
    • A23L3/3454—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals in the form of liquids or solids
    • A23L3/3463—Organic compounds; Microorganisms; Enzymes
    • A23L3/3571—Microorganisms; Enzymes
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
    • A01N63/30—Microbial fungi; Substances produced thereby or obtained therefrom
    • A01N63/32—Yeast
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23B—PRESERVING, e.g. BY CANNING, MEAT, FISH, EGGS, FRUIT, VEGETABLES, EDIBLE SEEDS; CHEMICAL RIPENING OF FRUIT OR VEGETABLES; THE PRESERVED, RIPENED, OR CANNED PRODUCTS
    • A23B4/00—General methods for preserving meat, sausages, fish or fish products
    • A23B4/12—Preserving with acids; Acid fermentation
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23B—PRESERVING, e.g. BY CANNING, MEAT, FISH, EGGS, FRUIT, VEGETABLES, EDIBLE SEEDS; CHEMICAL RIPENING OF FRUIT OR VEGETABLES; THE PRESERVED, RIPENED, OR CANNED PRODUCTS
    • A23B4/00—General methods for preserving meat, sausages, fish or fish products
    • A23B4/14—Preserving with chemicals not covered by groups A23B4/02 or A23B4/12
    • A23B4/18—Preserving with chemicals not covered by groups A23B4/02 or A23B4/12 in the form of liquids or solids
    • A23B4/20—Organic compounds; Microorganisms; Enzymes
    • A23B4/22—Microorganisms; Enzymes; Antibiotics
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/02—Separating microorganisms from their culture media
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/14—Fungi; Culture media therefor
    • C12N1/16—Yeasts; Culture media therefor
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs

Definitions

  • the invention relates to the field of biological antisepsis preservation and anti-oxidation, in particular to a preparation method of a fish bait preservative cultivated by bioengineering technology.
  • Fishing lures are a variety of special fishing bait, the pursuit of harmless, nutritious, the key is to play the role of poly fish and attract fish bites.
  • the original wild fish has a special liking for animal meat with high animal protein and natural fresh live bait.
  • the olfactory sensitivity of fish is hundreds of times higher than that of humans, especially for aquatic animals such as snails, shellfish, otters, shrimps, small fish, red worms, silkworms, seafood, etc.
  • Amino acids and riboflavin, niacin-like organisms are more interested, and because their generations are grown in water, these aquatic products are the main food source, and their genetics have been deeply imprinted, thinking that eating these foods is delicious. It is safe, so using these products as a bait is excellent.
  • aquatic products and meats used for fishing baits are not suitable for long-term storage at room temperature, and are often prone to decay, affecting the use effect, and are unsanitary and easy to spread many bacteria. Therefore, there is an urgent need for a good preservation effect on the market. It does not destroy the preservative of the original ingredients of the original fish bait.
  • the invention develops a preparation method of a fish bait preservative cultivated by bioengineering technology in order to solve the problem that the existing fishing baits made of fresh meat, aquatic meat and whole are not easy to store at room temperature.
  • the method for preparing the preservative is simple, efficient, and low in cost, and can avoid the fresh bait (dry and wet state) from being spoiled and deteriorated during the normal temperature preservation process, and the dry consumption is caused by blocks, mites, powder flakes, pastes, pastes. It is more convenient to store and use the bait such as liquid.
  • the present invention provides a preparation method of a fish bait preservative cultivated by bioengineering technology, comprising the following steps:
  • the distillation time is guaranteed to be more than 40 minutes to kill the bacteria and retain the effective bacteria such as heat-resistant koji yeast and lactic acid bacteria.
  • the above-mentioned distilled koji temperature is cooled to below 40 ° C, and then placed in a plastic bag for vacuum packaging, the volume of the plastic bag is between 500-1000 g, and the anaerobic sterilization process is performed, and the anaerobic yeast is further purified.
  • lactic acid bacteria
  • the steamed and stuffy above materials are out of the cage and spread to a temperature below 45 °C;
  • the parent species in the second step is incorporated, and the parent species accounts for 1%-10% of the total proportion, and then fully mixed;
  • the mixing material in the above steps is dry, add the steamer bottom water to dilute.
  • the temperature of the bottom water of the steamer is required to be below 50 °C, so that the material is in a wet state, and it is better to have water droplets in the hand holding mass.
  • the specific water content should be above 50%;
  • the above-mentioned uniformly mixed raw materials are placed in a container with good sealing performance for sealing fermentation.
  • the volume of the container is between 50 and 500 kg.
  • Total number of bacteria according to GB
  • the total number of bacteria detected by the test method 4789.2-2010 should be between 9.0x105-8.2x108cfu/g;
  • the total number of yeasts detected by the method of 4789.15-2010 should be between 4.8x103-5.0x104cfu/g, according to GB.
  • the total number of yeasts detected by the method of 4789.35-2010 should be between 1.2x107 and 1.1x108 cfu/g;
  • the qualified fermented material can be packaged after treatment.
  • the fermented grain in the third step is according to GB
  • the total number of yeasts detected by the method of 4789.15-2010 should be between 2.7x104-3.1x105cfu/g, according to GB.
  • the total number of yeasts detected by the method of 4789.35-2010 should be between 2.7x107-8.1x107cfu/g, according to GB/T
  • the glucose content detected by the detection method of 22221-2008 should be between 0-0.4%.
  • the fructose content detected according to the test method of GB/T 22221-2008 should be between 0-0.4%.
  • the qualified fermented product can be made into a paste, a water, a paste or a powder.
  • the fish bait preservative cultivated by bioengineering technology partially restores the underwater natural ecosystem, not only can maintain the original state (shape, taste, color) of the bait, but also can effectively control other harmful bacteria under normal temperature conditions. Breeding, maintaining the activity of animal and vegetable protein bait, minimizing the degradation rate of plant protein and various meats, and long-term storage will also have some fat, protein is converted into fish, especially loved amino acids, riboflavin , dimethyl disulfide, dimethyl trisulfide and partial esters, ethers, phenolic compounds, just to play an excellent role in attracting food, does not affect the use, but improve the effectiveness, for the development and preservation of bait Broad prospects.
  • the preparation method of the fish bait preservative cultivated by the bioengineering technology of the invention utilizes the characteristics of the yeast, the lactic acid bacteria and the enzyme which are both facultative. Specifically, the combined strains of yeast, lactic acid bacteria and their enzymes optimized by high-quality koji yeast are used in the medium made of high-quality starch as the main material, vegetable protein and plant fiber as auxiliary materials, and water gelatinization.
  • Oxygen-controlled fermentation cultivation artificial regulation of a-amylase activity, through a combination of bacteria, enzymes and biocatalysts, through a series of complex biochemical reactions, ultimately to maintain the activity of animal and plant tissue cells and their defects
  • the polysaccharides and hyaluronic acid components which are repaired by the cells, form a paste, water, paste or powder-like bait preservative with unique preservation, antiseptic and anti-oxidation effects.
  • a preparation method of a fish bait preservative cultivated by bioengineering technology comprising:
  • step 1 The high-quality koji yeast is steamed at 60 ° C for 40-45 minutes, then spread out to 40 ° C, and it is vacuum-packed in a plastic bag with a mass unit of 500 to 1000 g, and is kept at room temperature for 15-30 days.
  • the white yeast hyphae are formed on the edge of the plastic bag, the sample is tested and the test can be used as the parent species.
  • Step 2 Mix the high-quality demineralized gray glutinous rice and glutinous rice that have been tested and screened cleanly in a ratio of 4:1, then add water in a ratio of 1:1, steam for 40 minutes in the cage, steam for 15 minutes after steaming, then Power off and turn off the fire, continue to suffocate for 40 minutes, and spread out to the air below 45 °C; the mature mother in the first step is mixed with the above materials in a ratio of 1%, and the water content is adjusted with the bottom water of the steamer at 50 °C. More than 50%, and then into the large-scale sealing performance of the fermenter for oxygen-controlled room temperature fermentation, and appropriate decompression of the fermenter in a timely manner, to maintain normal pressure.
  • Step 3 After about 15-30 days, when the fermented material in the fermenter is observed or sampled with white transparent liquid, no gas is produced, no need to decompress, sample and test, after the test meets the standard, the fermentation will be carried out. Material paste, liquid preparation, powder preparation for use.
  • a preparation method of a fish bait preservative cultivated by bioengineering technology comprising:
  • Step 1 The high-quality rice wine koji yeast is continuously heated by steam at 60 ° C for 50 minutes, then spread out to 35 ° C, and then it is vacuum-packed in a plastic bag with a mass unit of 500 to 1000 g, and is kept under normal temperature environment. 30 days, when the white yeast hyphae are formed on the edge of the plastic bag, the sample is tested and the test can be used as the parent species.
  • Step 2 Mix the gray-white glutinous rice and glutinous rice with good quality and good quality, and remove the endothelium.
  • the mixture is mixed in a ratio of 1:1, then added to the water in a ratio of 1:1, steamed for 40 minutes in the cage, and steamed for 15 minutes after steaming. Then turn off the power and turn off the fire, continue to suffocate for 50 minutes, and spread out to 40 °C;
  • the mature mother in the first step is mixed with the above raw materials in a proportion of 5%, and the water content is adjusted with the bottom water of the steamer at 45 °C. More than 50%, and then into the large-scale sealing performance of the fermenter for oxygen-controlled room temperature fermentation, and appropriate decompression of the fermenter in a timely manner, to maintain normal pressure.
  • Step 3 After about 15-30 days, when the fermented material in the fermenter is observed or sampled with white transparent liquid, no gas is produced, no need to decompress, sample and test, after the test meets the standard, the fermentation will be carried out. Material paste, liquid preparation, powder preparation for use.
  • a preparation method of a fish bait preservative cultivated by bioengineering technology comprising:
  • step 1 The high-quality koji yeast is continuously heated by steam at 60 ° C for 40 minutes, taken out and stored at room temperature for 24 hours, and then it is vacuum-packed in a plastic bag at a mass unit of 500 to 1000 g, and is kept at a normal temperature for 15-30 days.
  • the white yeast hyphae are formed on the edge of the plastic bag, the sample is tested and the test can be used as the parent species.
  • Step 2 Mix the gray-white glutinous rice and the rice with good quality and good descreening, and mix it in a ratio of 4:1, then add the water in a ratio of 1:1, steam for 40 minutes, steam for 15 minutes after steaming. Then turn off the power and turn off the fire, continue to suffocate for 45 minutes, and spread out to 40 °C; the mature mother in the first step is mixed with the above materials in a proportion of 1%, and the water content is adjusted with the bottom water of the steamer at 40 °C. More than 50%, and then into the large-scale sealing performance of the fermenter for oxygen-controlled room temperature fermentation, and appropriate decompression of the fermenter in a timely manner, to maintain normal pressure.
  • Step 3 After about 15-30 days, when the fermented material in the fermenter is clear by observation or sampling with a white transparent liquid, and the pressure gauge shows that there is no gas output, no need to decompress, sample test, test After passing the standard, the fermented product is made into a paste, liquid, and powder for use.
  • the detection standard of the fermented grain is: according to GB
  • the total number of yeasts detected by the method of 4789.15-2010 should be between 2.7x104-3.1x105cfu/g, according to GB.
  • the total number of yeasts detected by the method of 4789.35-2010 should be between 2.7x107-8.1x107cfu/g, according to GB/T
  • the glucose content detected by the detection method of 22221-2008 should be between 0-0.4%.
  • the fructose content detected according to the test method of GB/T 22221-2008 should be between 0-0.4%.
  • the final produced fermentation product detection standard is:
  • Total number of bacteria according to GB
  • the total number of bacteria detected by the test method 4789.2-2010 should be between 9.0x105-8.2x108cfu/g;
  • the total number of yeasts detected by the method of 4789.15-2010 should be between 4.8x103-5.0x104cfu/g, according to GB.
  • the total number of yeasts detected by the method of 4789.35-2010 should be between 1.2x107 and 1.1x108 cfu/g;
  • the preservative should be wrapped or mixed with the fresh-keeping material. Then, the fresh-keeping material is wrapped with plastic wrap or sealed in a sealed container to allow the preservative to be in full contact with the fresh-keeping material.
  • the active factor polysaccharide and hyaluronic acid are rapidly introduced into the cells of the fresh-keeping material as an exogenous stress active agent, thereby ensuring cell humidity and nutrient stability, and not in a short period of time.
  • the yeast and lactic acid bacteria in the preservative will be closely combined with the fresh-keeping material, and colonized and bred on the cell enzyme to form a uniform and strict protective film, which not only blocks other harmful bacteria.
  • the preservative will continue to produce the polysaccharides, hyaluronic acid and other active agents required for the preservation of fresh materials, to ensure the long-term freshness of the preservation of fresh materials and the nutrient metabolism of the cell parts.
  • the present invention will greatly maintain the sensory, feel, and taste properties of the fresh-keeping materials, and with a small amount of protein and fat over time.
  • active enzymes in the preservative Under the decomposition of active enzymes in the preservative, the amino acids, riboflavin, dimethyl disulfide, dimethyl trisulfide and partial esters, ethers and phenolic compounds that fish are particularly loved are formed.
  • a good lure effect not only does not affect the use, but also improves the effectiveness of the bait.
  • the receptor when used as a preservative, after the above steps are carried out, the receptor (preserved material) may be affected by complicated factors such as drying, water immersion, drug immersion, water loss, dryness, water swelling, and tissue cells. There is a phenomenon of water loss and shrinkage. At this time, the tissue cells of the fresh-keeping substance temporarily appear dormant, but as long as the temperature is not lower than minus 40 ° C and not higher than 45 ° C, the polysaccharide and hyaluronic acid in the recipient tissue cells will maintain a certain activity of the receptor, and will not Corruption and dry consumption have occurred. When the appropriate conditions are met, the recipient cells will rapidly swell or remove excess water and return to normal tissue state for long-term preservation.
  • the qualified fermented product is dried by hot air of 50 ° C or less or naturally dried, and when the moisture content thereof is less than 30%, it is crushed into a powder, and is passed below 80 mesh. Screen (can also be sieved to ensure that the cell structure is not destroyed); then vacuum packed with plastic bags (plastic bag volume unit 500g-1000g), the main purpose is to kill the bacteria infection, to Maintain effective bacterial purity and effective active ingredients without loss.
  • the above-mentioned dried powder is mixed with the material to be treated in a ratio of 0.2% to 20%, and is preserved at normal temperature and normal pressure to have an antioxidant effect. Materials containing this preservative can also be stored in a freezer, vacuum, etc. without being affected.
  • the use of the invention as an antioxidant can solve the problems of ashing, blackening, lightening, odor and other oxidative deterioration such as fish meal, shrimp powder, dried plant products and dried meat products, achieving long-term natural preservation and slowing the loss of nutrient components. effect.

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Abstract

一种鱼饵保鲜剂的制备方法，包括母种的选种、冷却、发酵谷物的蒸馏备料、接种、检测及包装。该方法采用优质酒曲酵母优化出的酵母菌、乳酸菌及其酶等联合菌株，在以优质淀粉为主要材料、植物蛋白和植物纤维为辅助材料与水加热糊化做出的培养基中进行控氧发酵培植，人为调控α淀粉酶的活性，从而在其菌、酶及生物催化剂的共同作用下，通过一系列复杂的生化反应，最终生成具有保持动植物组织细胞活性并对其缺损细胞进行修复作用的多糖类和透明质酸等成分，形成具有保鲜、防腐、抗氧化效果的鱼饵保鲜剂。

Description

一种采用生物工程技术培植的鱼饵保鲜剂的制备方法 技术领域
本发明涉及生物防腐保鲜抗氧化领域，具体地说，涉及一种采用生物工程技术培植的鱼饵保鲜剂的制备方法。
 背景技术
鱼饵是多种特殊的钓鱼用饵料，追求的是无害、营养，关键是能够起到聚鱼和引诱鱼咬钩的作用。而原生态的野生鱼类对动物蛋白高的动物鲜肉及天然鲜活动物性饵料情有独钟。鱼类的嗅觉敏感度是人类的数百倍，尤其对水生动物，如螺、贝、水蚯蚓、虾、小鱼、红虫、沙蚕、海产品等，鱼类对它们其中含有的大量多种氨基酸及核黄素、烟酸的生物更加感兴趣，并且由于其世代生长在水中，以这些水产为主要食物源，其遗传基因里已深深的打上烙印，认为觅食这类食物既可口又安全，因此采用这些产品作为鱼饵效果极佳。
 技术问题
但做鱼饵用的水产品、肉类等却不宜在常温下长期保存，往往容易腐变，影响使用效果，而且不卫生，容易传播许多细菌，因此市场上急需一款具有良好的保鲜效果，而又不破坏原有鱼饵料原有成分的保鲜剂。
 技术解决方案
本发明为了解决现有以鲜肉、水产肉类体及整体做的鱼饵在常温下不易保存的问题，开发了一种采用生物工程技术培植的鱼饵保鲜剂的制备方法。该保鲜剂制作方法简单、高效、成本低廉、能避免新鲜鱼饵（干湿态）在常温保存过程中不易腐败变质、干耗，使块类、糜类、粉片状、浆膏状、糊状、液态等鱼饵的保管及使用更方便。
为解决上述问题，本发明提供一种采用生物工程技术培植的鱼饵保鲜剂的制备方法，包括以下步骤：
一、选种：
选取优质的酒曲酵母，将其放入60℃的环境温度下进行蒸馏，蒸馏时间保证在40分钟以上，以杀灭杂菌，保留其中耐热的酒曲酵母菌、乳酸菌等类型的有效菌；
二、冷却：
①. 将上述蒸馏后的酒曲温度冷却至40℃以下，然后装入塑料袋进行真空包装，塑料袋的容积在500-1000g之间，进行无氧灭菌程序，同时进一步纯化具有兼氧性的酵母菌及乳酸菌；
②.将塑料袋在常温环境下放置15-30天，当包装袋边缘有白色酵母菌丝出现时取样检测，检测合格则表示母种的制备完成；
三、备料：
①. 选取发酵谷物为去除掉内皮的灰白色高粱米和糯米或者去除掉内皮的灰白色高粱米和大米，筛选无杂质并检测合格后，按4:1的比例混合均匀，再按1:1的比例加入清水，上笼蒸馏40-50分钟，上汽后再延时10-15分钟，然后断电关火，继续闷40-50分钟；
②. 将蒸熟闷透的上述原料出笼、摊晾，使其温度降至45℃以下；
四、接种：
①. 在上述步骤三中的原料中掺入步骤二中的母种，母种含量占整体比例的1%-10%，然后充分拌匀；
②.上述步骤中的拌匀的材料若偏干，可添加蒸笼底水进行稀释，蒸笼底水的温度要求在50℃以下，使材料呈现湿润状态，以手握料团有水滴出为宜，具体含水量应在50%以上；
③.将上述混合均匀后的原材料放入密封性能良好的容器中进行密封发酵，容器的容积在50-500kg之间，密封期间需注意容器的排气及减压，保持常压状态，然后在常温下自然发酵15天以上，得到发酵物，再以碳酸氢钠、过氧化钙或氧化三甲胺将发酵物中和至PH值4-6.5即可；
五、 检测
①. 细菌总数：根据GB 4789.2-2010检测方法检测出的细菌总数应在9.0x105-8.2x108cfu/g之间；
②. 有效菌：根据GB 4789.15-2010检测方法检测出的酵母菌总数应在4.8x103-5.0x104cfu/g之间，根据GB 4789.35-2010检测方法检测出的酵母菌总数应在1.2x107-1.1x108cfu/g之间；
③. 根据GB 4789.15-2010检测方法检测中温α-淀粉酶活力范围应在18.0-38.1u/g之间。
六、包装：将检测合格的发酵物处理后包装即可。
优选地，所述步骤三中发酵谷物根据GB 4789.15-2010检测方法检测出的酵母菌总数应在2.7x104-3.1x105cfu/g之间，根据GB 4789.35-2010检测方法检测出的酵母菌总数应在2.7x107-8.1x107cfu/g之间，根据GB/T 22221-2008检测方法检测出的葡萄糖含量应在0-0.4%之间，根据GB/T 22221-2008检测方法检测出的果糖含量应在0-0.4%之间。
优选地，所述检测合格的发酵物可制成糊状、水状、膏状或者粉剂状。
因为在水体下面除了活的生物外，还会有死亡的动物尸体及淹没的庄稼地里的粮食、草种、瓜果、蚯蚓及各种生物体，时间一长，在水体及水下的土壤中的酵母菌、乳酸菌等会在这些腐败的生物上面繁殖，分解它们所含的糖类、蛋白质等，进而生成新的成分诸如乙醇、酚类、醚类、酯类等挥发性和水溶性极强的物质，这些成分会随着水流不断的扩散出去，鱼类受其遗传基因及灵敏嗅觉的识别指令，就会主动寻找这些味源，因此这类鱼饵更易引诱鱼咬钩。
 有益效果
本发明一种采用生物工程技术培植的鱼饵保鲜剂，部分还原水下自然生态系统，不仅能够保持鱼饵的原始状态（形、味、色），而且在常温情况下，可以有效的控制其它有害细菌滋生，保持动植物蛋白饵料的活性，最大限度的减缓植物蛋白及多种肉类的降解速度，并且长期存放还会有部分的脂肪、蛋白被转化成鱼类特别爱吃的氨基酸、核黄素、二甲基二硫、二甲基三硫及部分酯、醚、酚类化合物，正好起到极好的诱食作用，不影响使用，反而提高了有效性，对于饵类的开发及保存有着广阔的前景。
本发明一种采用生物工程技术培植的鱼饵保鲜剂的制备方法利用的是均具兼氧性的酵母菌、乳酸菌及其酶的特性。具体的是采用优质酒曲酵母优化出的酵母菌、乳酸菌及其酶等联合菌株，经在以优质淀粉为主要材料，植物蛋白和植物纤维为辅助材料，与水加热糊化做出的培养基中进行控氧发酵培植，人为调控a-淀粉酶的活性，从而在其菌、酶及生物催化剂的共同作用下，通过一系列复杂的生化反应，最终生成具有保持动植物组织细胞活性并对其缺损细胞进行修复作用的多糖类和透明质酸等成分，形成具有独到的保鲜、防腐、抗氧化效果的糊状、水状、膏状或者粉剂状的鱼饵保鲜剂。
 附图说明
本发明的最佳实施方式
本发明的实施方式
为了使本领域的技术人员更好的理解本发明的技术方案，下面结合具体实施例对本发明的技术方案进行清楚、完整地描述。
实施例1：
一种采用生物工程技术培植的鱼饵保鲜剂的制备方法，包括：
第1步： 将优质酒曲酵母经60℃蒸汽蒸40-45分钟，然后出笼摊开晾到40℃，将其以500到1000g的质量单位装入塑料袋真空包装，在常温环境下待15-30天，待塑料袋边缘有白色酵母菌丝生成时，抽样检测，检测达标后即可作为母种。
第2步：将检测合格筛选干净的优质去内皮灰白色高粱米和糯米按4:1比例混合均匀，再按1:1的比例加入清水，上笼蒸40分钟，上汽后再蒸15分钟，然后断电关火，继续闷40分钟，出笼摊开晾至45℃以下；将第一步中成熟的母种按1%的比例与上述原料充分混合均匀，用50℃的蒸笼底水调整含水量50%以上，再将其装入大型密封性能良好的发酵罐中进行控氧常温发酵，并适当适时对发酵罐进行排气减压，保持常压。
第3步：约15-30天后，当发酵罐内的发酵物通过观察或取样有白色透明液体晰出、无气体产出、不再需要减压时，取样化验，检测合格达标后，将发酵物制浆膏、制液、制粉待用。
实施例2：
一种采用生物工程技术培植的鱼饵保鲜剂的制备方法，包括：
第1步：将优质米酒酒曲酵母经60℃蒸汽连续加热50分钟，然后出笼摊开晾到35℃，将其以500到1000g的质量单位装入塑料袋真空包装，在常温环境下待15-30天，待塑料袋边缘有白色酵母菌丝生成时，抽样检测，检测达标后即可作为母种。
第2步：将检测合格筛选干净的优质脱去内皮的灰白色高粱米和糯米按4:1比例混合，再按1:1的比例加入清水，上笼蒸40分钟，上汽后再蒸15分钟，然后断电关火，继续闷50分钟，出笼摊开晾至40℃；将第一步中成熟的母种按5%的比例与上述原料充分混合均匀，用45℃的蒸笼底水调整含水量50%以上，再将其装入大型密封性能良好的发酵罐中进行控氧常温发酵，并适当适时对发酵罐进行排气减压，保持常压。
第3步：约15-30天后，当发酵罐内的发酵物通过观察或取样有白色透明液体晰出、无气体产出、不再需要减压时，取样化验，检测合格达标后，将发酵物制浆膏、制液、制粉待用。
实施例3：
一种采用生物工程技术培植的鱼饵保鲜剂的制备方法，包括：
第1步: 将优质酒曲酵母经60℃蒸汽连续加热40分钟，将其取出在常温状态下存放24小时，然后将其以500到1000g的质量单位装入塑料袋真空包装，在常温环境下待15-30天，待塑料袋边缘有白色酵母菌丝生成时，抽样检测，检测达标后即可作为母种。
第2步：将检测合格筛选干净的优质脱去内皮的灰白色高粱米和大米按4:1比例混合，再按1:1的比例加入清水，上笼蒸40分钟，上汽后再蒸15分钟，然后断电关火，继续闷45分钟，出笼摊开晾至40℃；将第一步中成熟的母种按1%的比例与上述原料充分混合均匀，用40℃的蒸笼底水调整含水量50%以上，再将其装入大型密封性能良好的发酵罐中进行控氧常温发酵，并适当适时对发酵罐进行排气减压，保持常压。
第3步：约15-30天后，当发酵罐内的发酵物通过观察或取样有白色透明液体晰出，而压力表显示不再有气体产出、不再需要减压时，取样化验，检测合格达标后，将发酵物制浆膏、制液、制粉待用。
在本发明中，发酵谷物的检测标准为：根据GB 4789.15-2010检测方法检测出的酵母菌总数应在2.7x104-3.1x105cfu/g之间，根据GB 4789.35-2010检测方法检测出的酵母菌总数应在2.7x107-8.1x107cfu/g之间，根据GB/T 22221-2008检测方法检测出的葡萄糖含量应在0-0.4%之间，根据GB/T 22221-2008检测方法检测出的果糖含量应在0-0.4%之间。
在本发明中，最终生成的发酵物检测标准为：
①. 细菌总数：根据GB 4789.2-2010检测方法检测出的细菌总数应在9.0x105-8.2x108cfu/g之间；
②. 有效菌：根据GB 4789.15-2010检测方法检测出的酵母菌总数应在4.8x103-5.0x104cfu/g之间，根据GB 4789.35-2010检测方法检测出的酵母菌总数应在1.2x107-1.1x108cfu/g之间；
③. 根据GB 4789.15-2010检测方法检测中温α-淀粉酶活力范围应在18.0-38.1u/g之间；
本发明用作保鲜剂使用时的具体操作方法为：
①、将新鲜的肉块或海鲜、水产类动物整体清洗干净后，加入两倍于原料体积的酒对其进行清洗、浸泡，作初步清洗及杀菌，其中浸泡时间为10-20分钟，酒的度数范围为40-65度，最终溶出部分原料自身的活性酶，为接种的菌、酶的占位、着床、定植做准备。
②、将上述①中浸泡后的原料沥干，加入两倍于被保鲜材料质量或体积的保鲜剂，保鲜剂要大体将被保鲜材料包裹或混合覆盖。然后将被保鲜材料用保鲜膜包裹或者将其放入密封容器内密封，让保鲜剂与被保鲜材料充分接触。
该保鲜剂经上述步骤应用到被保鲜材料上后，其中的活性因子多糖、透明质酸作为外源性应激活性剂迅速进入被保鲜材料的细胞内，保证细胞湿度及养分稳定，短期内不至凋亡，同时保鲜剂中的酵母菌、乳酸菌也会很快与被保鲜材料紧密结合，并在其细胞酶上定植、繁育，形成一层均匀、严密的保护膜，不仅阻断其它有害细菌的入侵其中，防止腐败，另外该保鲜剂还会继续生成被保鲜材料需要的多糖、透明质酸等活性剂，保证被保鲜材料的长期鲜活及细胞部分的营养代谢。
一般情况下，常温常压下在6个月之内，本发明会极大地保持被保鲜材料的感官、手感、味型的良好性能，并且随着时间的延长，还会有少量的蛋白、脂肪在该保鲜剂内活性菌酶的分解下，形成鱼类特别爱吃的氨基酸、核黄素、二甲基二硫、二甲基三硫及部分酯、醚、酚类化合物，正好起到了极好的诱食效果，不仅不影响使用，反而提高了鱼饵的有效性。
本发明用作防腐剂使用时的具体操作方法为：
①、将新鲜的肉块或海鲜、水产类动物整体清洗干净后，加入两倍以上于原料体积的酒对其进行清洗、浸泡，作初步清洗及杀菌，其中浸泡时间为10-20分钟，酒的度数范围为40-65度，最终溶出部分原料自身的活性酶，为接种的菌、酶的占位、着床、定植做准备。
②、将上述①中浸泡后的原料沥干，加入两倍以上于被处理材料质量或体积的保鲜剂，保鲜剂要大体将被处理材料包裹或混合。然后将被处理材料用保鲜膜包裹或者将其放入密封容器内密封，让保鲜剂与被处理材料充分接触。
本发明作为防腐剂使用时，按上述步骤实施以后，受体（被保鲜材料）可能会因遇到干燥、水浸、药浸等复杂因素的影响，出现失水干燥、遇水膨胀、组织细胞出现失水缩水等现象。这时被保鲜物质的组织细胞暂时呈现休眠状态，但只要温度不低于零下40℃、不高于45℃，受体组织细胞中的多糖、透明质酸会保持受体一定的活性，不会出现腐败、干耗现象。当遇到合适的条件，受体细胞会迅速吸水膨胀或脱去多余的水分，恢复正常组织状态，以达到长期防腐的目的。
本发明用作抗氧化剂使用时的具体操作方法为：
①、本发明作为抗氧化剂使用时，首先将检测合格的发酵物通过50℃以下的热风干燥或自然干燥，当其水分含量达到30%以下时，将其碾碎成粉状，过80目以下的筛（也可以不过筛，以保证菌体组织结构不被破坏）；然后使用塑料袋对其进行真空包装（塑料袋容积单位为500g-1000g），其主要目的是杀灭杂菌感染，以保持有效菌纯度及有效活性成分不受损失。
②、然后将上述干燥后的粉剂与被处理材料按0.2%-20%的比例混合，在常温常压保存即可起到抗氧化作用。含有本防腐剂的材料也可以进行冷冻储藏、真空保存等操作，不会受到影响。
本发明用作抗氧化剂使用可以解决诸如鱼粉、虾粉、植物干品及肉类干品的变灰、变黑、变淡、变味等氧化变质的问题，达到长期自然保存，减缓营养成分耗损的效果。
 工业实用性
本发明不局限于上述的优选实施方式，任何人应该得知在本发明的启示下做出的结构变化，凡是与本发明具有相同或者相近似的技术方案，均属于本发明的保护范围。
 序列表自由内容

Claims (3)

  1. 一种采用生物工程技术培植的鱼饵保鲜剂的制备方法，其特征在于，包括以下步骤：
    一、选种：
    选取优质的酒曲酵母，将其放入60℃的环境温度下进行蒸馏，蒸馏时间保证在40分钟以上，以杀灭杂菌，保留其中耐热的酒曲酵母菌、及乳酸菌这类有效菌；
    二、冷却：
    ①. 将上述蒸馏后的酒曲温度冷却至40℃以下，然后装入塑料袋进行真空包装，塑料袋的容积在500-1000g之间，进行无氧灭菌程序，同时进一步纯化具有兼氧性的酵母菌及乳酸菌；
    ②.将塑料袋在常温环境下放置15-30天，当包装袋边缘有白色酵母菌丝出现时取样检测，检测合格则表示母种的制备完成；
    三、备料：
    ①. 选取发酵谷物为去除掉内皮的灰白色高粱米和糯米或者去除掉内皮的灰白色高粱米和大米，筛选无杂质并检测合格后，按4:1的比例混合均匀，再按1:1的比例加入清水，上笼蒸馏40-50分钟，上汽后再延时10-15分钟，然后断电关火，继续闷40-50分钟；
    ②. 将蒸熟闷透的上述原料出笼、摊晾，使其温度降至45℃以下；
    四、接种：
    ①. 在上述步骤三中的原料中掺入步骤二中的母种，母种含量占整体比例的1%-10%，然后充分拌匀；
    ②.上述步骤中的拌匀的材料若偏干，可添加蒸笼底水进行稀释，蒸笼底水的温度要求在50℃以下，使材料呈现湿润状态，以手握料团有水滴出为宜，具体含水量应在50%以上；
    ③. 将上述混合均匀后的原材料放入密封性能良好的容器中进行密封发酵，容器的容积在50-500kg之间，密封期间需注意容器的排气及减压，保持常压状态，然后在常温下自然发酵15天以上，得到发酵物，再以碳酸氢钠、过氧化钙或氧化三甲胺将发酵物中和至PH值4-6.5即可；
    五、检测
    ①. 细菌总数：根据GB 4789.2-2010检测方法检测出的细菌总数应在9.0x105-8.2x108cfu/g之间；
    ②. 有效菌：根据GB 4789.15-2010检测方法检测出的酵母菌总数应在4.8x103-5.0x104cfu/g之间，根据GB 4789.35-2010检测方法检测出的酵母菌总数应在1.2x107-1.1x108cfu/g之间；
    ③. 根据GB 4789.15-2010检测方法检测中温α-淀粉酶活力范围应在18.0-38.1u/g之间；
    六、包装：将检测合格的发酵物处理后包装即可。
  2. 根据权利要求1所述的一种采用生物工程技术培植的鱼饵保鲜剂的制备方法，其特征在于：所述步骤三中发酵谷物根据GB 4789.15-2010检测方法检测出的酵母菌总数应在2.7x104-3.1x105cfu/g之间，根据GB 4789.35-2010检测方法检测出的酵母菌总数应在2.7x107-8.1x107cfu/g之间，根据GB/T 22221-2008检测方法检测出的葡萄糖含量应在0-0.4%之间，根据GB/T 22221-2008检测方法检测出的果糖含量应在0-0.4%之间。
  3. 根据权利要求1所述的一种采用生物工程技术培植的鱼饵保鲜剂的制备方法，其特征在于：所述检测合格的发酵物可制成糊状、水状、膏状或者粉剂状。
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114468000A (zh) * 2021-12-24 2022-05-13 多麦(福建)食品有限公司 一种耐储存面包及其制备方法

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114456967B (zh) * 2021-10-14 2022-10-04 命之源(杭州)生物科技有限公司 一种酵母、乳酸菌联合菌、培育方法及其应用

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101724561A (zh) * 2008-10-15 2010-06-09 陈佩仁 生料黄酒曲的制备方法
CN102960570A (zh) * 2012-12-10 2013-03-13 北海市翰华生物技术有限公司 一种水产饲料的加工防腐方法
CN102972661A (zh) * 2012-12-17 2013-03-20 福建格林生物科技有限公司 高强耐水耐贮存凝胶状水产饲料
CN102972662A (zh) * 2012-12-17 2013-03-20 福建格林生物科技有限公司 膏状水产饲料

Family Cites Families (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5776523A (en) * 1996-12-27 1998-07-07 Axelrod; Herbert R. Method for preserving baits
CN1389162A (zh) * 2001-06-06 2003-01-08 王全根 一种无公害生物活性食物保鲜剂及其制备方法
CN101653168A (zh) * 2008-08-18 2010-02-24 邓卫欣 果菜生物保鲜剂
CN102583776B (zh) * 2012-02-29 2013-04-24 中国水产科学研究院淡水渔业研究中心 一种改良养殖水体环境的复合菌酶制剂及制备方法
CN103190470B (zh) * 2013-03-24 2015-09-09 新疆农垦科学院 一种肉类复合生物保鲜剂及其制备方法
CN104798749A (zh) * 2015-05-12 2015-07-29 湖北龙王恨渔具集团有限公司 一种生物发酵的鱼饵
CN106036198A (zh) * 2016-05-25 2016-10-26 大连工业大学 一种海参生物发酵饵料的制备方法

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101724561A (zh) * 2008-10-15 2010-06-09 陈佩仁 生料黄酒曲的制备方法
CN102960570A (zh) * 2012-12-10 2013-03-13 北海市翰华生物技术有限公司 一种水产饲料的加工防腐方法
CN102972661A (zh) * 2012-12-17 2013-03-20 福建格林生物科技有限公司 高强耐水耐贮存凝胶状水产饲料
CN102972662A (zh) * 2012-12-17 2013-03-20 福建格林生物科技有限公司 膏状水产饲料

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114468000A (zh) * 2021-12-24 2022-05-13 多麦(福建)食品有限公司 一种耐储存面包及其制备方法

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