阅读说明：本技术 一种利用超声波破碎微藻的处理方法 (Treatment method for crushing microalgae by using ultrasonic waves ) 是由 张彬 邓潜 朱昔恩 熊建华 谢达祥 赵永贞 陈田聪 陈晓汉 韦嫔媛 于 2019-09-24 设计创作，主要内容包括：本发明公开一种利用超声波破碎微藻的处理方法,将富集有微藻的浓缩藻液与PBS缓冲液混合后,置于超声发生仪中处理20-40min,得到长度为2-30μm的微藻；所述微藻的形态为聚合群体、丝状体、长条带状中的任意一种。本发明克服了传统物理机械破碎方法因产生切力不匀和过大会导致微藻细胞体完整性受损的技术缺陷,本方法不破坏藻体的细胞壁,只是将形态为聚合群体、丝状体、长条带状的微藻联生体进行细胞之间的破碎分离,使成列、成群或成簇的细胞群破碎成长度更短的细胞列或细胞群,甚至是破碎成单个藻体细胞,而藻细胞内营养成分得以有效保留,本方法获得的微藻破碎液,满足水产动物育苗早期的摄食口径和育苗早期的营养需求。(The invention discloses a processing method for crushing microalgae by using ultrasonic waves, which comprises the steps of mixing concentrated microalgae solution enriched with microalgae with PBS buffer solution, and then placing the mixture in an ultrasonic generator for processing for 20-40min to obtain microalgae with the length of 2-30 mu m; the form of the microalgae is any one of polymeric group, filament and strip. The invention overcomes the technical defect that the integrity of the microalgae cell body is damaged due to uneven and overlarge shearing force generated by the traditional physical mechanical crushing method, the method does not damage the cell wall of the microalgae, but only carries out crushing separation on the cells of the microalgae symbiont in the shapes of polymeric groups, filaments and strip bands, so that the cell groups in rows, groups or clusters are crushed into cell rows or cell groups with shorter growth degree, even into single algae cells, and the nutrient components in the algae cells are effectively reserved.)

1. A treatment method for crushing microalgae by using ultrasonic waves is characterized by comprising the following steps: mixing the concentrated microalgae solution enriched with microalgae with PBS buffer solution, and treating the mixture in an ultrasonic generator with ultrasonic power of 40-80W for 20-40min, wherein the temperature of the mixture is 24-35 deg.C and the treated microalgae length is 2-30 μm; the form of the microalgae is any one of polymeric group, filament and strip.

2. The method as claimed in claim 1, wherein the cell density of microalgae in the concentrated solution is not less than 1.0 x 10⁷one/mL.

3. The method as claimed in claim 1, wherein the cell density of the microalgae in the concentrated solution is 1.0-6.0×10⁷one/mL.

4. The method as claimed in claim 1, wherein when the concentrated algal solution is mixed with PBS buffer, 300 and 400ml PBS buffer is added to each gram of concentrated algal solution.

Technical Field

The invention relates to a processing method for crushing microalgae by using ultrasonic waves.

Background

Algae are the lowest phylum of the plant kingdom, and belong to the autotrophic eukaryotes because their cells contain chlorophyll or other pigments, are capable of photosynthesis, self-synthesize nutrients required, live independently and multiply and grow primarily by cell division, and because microalgae are small in size and difficult to see by the naked eye, they generally require microscopic observation. There are 2 thousands of microalgae, and the primary producers of microalgae are the major water ecosystems, accounting for 40%, and provide the primary productivity for marine organisms. The special growth environment makes it contain various bioactive substances, such as active protein, vitamins, minerals, carotenoids, microalgae polysaccharide, polyunsaturated fatty acids, etc.

With the breakthrough of research on biological characteristics and large-scale cultivation of various economic microalgae in a freshwater environment, the microalgae are widely applied to large-scale seedling and cultivation of aquatic livestock such as marine fishes, shellfishes, shrimps, crabs and the like, and become key biological baits for the survival and the failure of various aquatic livestock. At present, more than 20 kinds of microalgae are widely used in aquatic seedling culture internationally. The microalgae commonly used in the aquatic seedling culture industry in China mainly comprise: chaetoceros gracilis, Chaetoceros muelleri, Thalassiosira wessoeri, Thalassiosira pseudonana, Phaeodactylum tricornutum, Skeletonema costatum, Navicula sp, Oncomelania benthamoides (Coccoceis sp.) of the diatom species; isochrysis such as Isochrysis major (Isochrysis zhangjiangensis) and Isochrysis minor (Isochrysis galbana) of Chrysophytes; platymonas subcordiformis (Platymonas subcordiformis) of the green algae species, Chlorella vulgaris (Chlorella vulgaris), Coccidium pavonica (Oosystis borgi), Nannochloropsis oculata (Nannochloropsis oculata), and Spirulina platensis (Spirolinia platensis) of the blue algae species, and the like.

The marine microalgae has an important effect on the early development of larvae of seawater fishes, shellfishes, shrimps and crabs, particularly when the larvae of prawns are transformed from nauplii to flea larvae. Platymonas and Chrysophyta can maintain the development and molting metamorphosis of prawn and crab; feeding chlorella preferentially in a face plate larva stage of the variegated little clam planktonic larva stage; in the process that the planktonic larvae of the abalones grow to young abalones, the rapid growth stage can be entered by utilizing the easily digestible diatom cell inclusion; the skeletonema costatum is widely used for culturing prawn larvae and has obvious effect. In addition, bioactive substances such as antibiotics contained or generated by the microalgae can kill pathogenic bacteria in water and improve the disease resistance of cultured animals; meanwhile, the microalgae also has important effects on improving the water quality of the aquaculture water body and regulating and controlling the balance of the micro-ecological environment of the water body, the bait microalgae is put into the seedling culture water body, so that the substances such as ammonia nitrogen, nitrite and the like can be directly absorbed and utilized, and meanwhile, the oxygen released by photosynthesis can also promote the nitrification of microorganisms on the ammonia nitrogen and the nitrite. In the prawn culture water environment, the Boji oocystis and the Nannochloropsis are artificially introduced, so that the water quality of the culture water body can be improved, and the blood cell number, the serum protein content and the activities of phenol oxidase, superoxide dismutase, lysozyme and antibacterial enzyme of the prawns are obviously improved compared with those of a control group.

However, in order to adapt to the optimum feeding caliber of each metamorphosis stage of the prawn larva, it is necessary to crush the microalgae with a large length and then feed the crushed microalgae. How to crush the microalgae to a proper length, and maintain the cell integrity of the microalgae after crushing and retain the biological activity is a technical problem to be solved urgently by the technical personnel in the field.

At present, the crushing of microalgae is performed by breaking cells of microalgae by physical and mechanical methods, and then releasing bioactive substances in the cells for use after the cell walls of the microalgae are broken. The existing method for crushing the microalgae cells has the problems of complex operation, poor crushing effect and serious loss of bioactive components in the cells.

There has been no report on the disruption of microalgae in the form of polymeric colonies, filaments, long ribbons between cells without damaging the cell wall and affecting bioactive components in the cells.

Disclosure of Invention

The invention aims to overcome the defects in the prior art and provides a treatment method for crushing microalgae by using ultrasonic waves.

In order to achieve the purpose, the technical scheme adopted by the invention is as follows:

the treatment method for crushing microalgae by using ultrasonic waves comprises the following steps: mixing the concentrated microalgae solution enriched with microalgae with PBS buffer solution, and treating the mixture in an ultrasonic generator with ultrasonic power of 40-80W for 20-40min, wherein the temperature of the mixture is 24-35 deg.C and the treated microalgae length is 2-30 μm; the form of the microalgae is any one of polymeric group, filament and strip.

Preferably, the cell density of the microalgae in the concentrated algae solution is not less than 1.0 x 10⁷one/mL.

Preferably, the cell density of the microalgae in the concentrated algae solution is 1.0-6.0 × 10⁷one/mL.

Preferably, when the concentrated algae solution is mixed with the PBS buffer solution, 300 and 400mLPBS buffer solution is added into each gram of the concentrated algae solution.

Preferably, the concentrated algal solution is algal somatic cells collected by concentrating a microalgal culture solution subjected to aseptic culture by centrifugation or the like.

Preferably, the rotation speed is 5000-.

Preferably, the microalgae in the form of long-strip ribbons is any one of the skeletonema.

Preferably, the microalgae in the form of a polymeric population or a filament is any one of the genera spirulina.

Due to the adoption of the technical scheme, the invention has the following beneficial effects:

1. the invention overcomes the technical defect that microalgae cell is damaged by crushing microalgae, realizes the crushing of microalgae in the form of polymeric group, filament and strip among cells, retains bioactive components in the cells, avoids the loss of bioactive components, and retains the nutritional components of the microalgae as far as possible.

2. In the process of crushing the microalgae, the protective agent is added and uniformly distributed around the microalgae particles to form an isolation barrier, so that the contact of oxygen and microalgae cell bodies is reduced, the active ingredients of the microalgae are protected from being oxidized and decomposed, and the bioactive ingredients of the microalgae are further ensured not to be oxidized and damaged.

3. According to the invention, through ultrasonic treatment, factors such as ultrasonic treatment time, ultrasonic treatment power, treatment temperature, algae liquid concentration and the like are subjected to multiple experimental screening, a set of technical process with high crushing efficiency and good stability is optimized, and the effective crushing rate can reach more than 95%.

4. The invention does not destroy the cell wall of the algae, but only breaks and separates the microalgae intergrowth in the form of polymeric group, filament and strip to make the cell group in line, group or cluster break into cell line or cell group with shorter growth degree, even break into single algae cell, and the nutrition content in the algae cell can be effectively preserved.

Drawings

FIG. 1 is a micrograph of Spirulina platensis algae at different sonication times.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention will be described in further detail below with reference to preferred embodiments. It should be noted, however, that the numerous details set forth in the description are merely for the purpose of providing the reader with a thorough understanding of one or more aspects of the present invention, which may be practiced without these specific details.