CN113813292B - Marine protozoan armillar extract with antibacterial activity - Google Patents
Marine protozoan armillar extract with antibacterial activity Download PDFInfo
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- 239000000284 extract Substances 0.000 title claims abstract description 56
- 230000000844 anti-bacterial effect Effects 0.000 title claims abstract description 27
- CPLXHLVBOLITMK-UHFFFAOYSA-N Magnesium oxide Chemical compound [Mg]=O CPLXHLVBOLITMK-UHFFFAOYSA-N 0.000 claims abstract description 42
- 235000012245 magnesium oxide Nutrition 0.000 claims abstract description 21
- 239000000395 magnesium oxide Substances 0.000 claims abstract description 21
- 241000863480 Vinca Species 0.000 claims abstract description 14
- 238000002360 preparation method Methods 0.000 claims abstract description 13
- 241000607598 Vibrio Species 0.000 claims abstract description 12
- 238000003113 dilution method Methods 0.000 claims abstract description 11
- RJQXTJLFIWVMTO-TYNCELHUSA-N Methicillin Chemical compound COC1=CC=CC(OC)=C1C(=O)N[C@@H]1C(=O)N2[C@@H](C(O)=O)C(C)(C)S[C@@H]21 RJQXTJLFIWVMTO-TYNCELHUSA-N 0.000 claims abstract description 6
- 241000700141 Rotifera Species 0.000 claims abstract description 6
- 241000191967 Staphylococcus aureus Species 0.000 claims abstract description 6
- 241000607265 Vibrio vulnificus Species 0.000 claims abstract description 6
- 229960003085 meticillin Drugs 0.000 claims abstract description 6
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 36
- 230000003321 amplification Effects 0.000 claims description 6
- 244000005700 microbiome Species 0.000 claims description 6
- 238000003199 nucleic acid amplification method Methods 0.000 claims description 6
- 239000013049 sediment Substances 0.000 claims description 6
- 241000255967 Helicoverpa zea Species 0.000 claims description 5
- 241000238631 Hexapoda Species 0.000 claims description 5
- 244000000010 microbial pathogen Species 0.000 claims description 5
- 239000001963 growth medium Substances 0.000 claims description 4
- 238000009630 liquid culture Methods 0.000 claims description 4
- 241001052560 Thallis Species 0.000 claims description 3
- 238000005119 centrifugation Methods 0.000 claims description 3
- 230000005764 inhibitory process Effects 0.000 claims description 3
- 239000006228 supernatant Substances 0.000 claims description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 3
- 239000006185 dispersion Substances 0.000 claims description 2
- 238000000605 extraction Methods 0.000 claims 1
- 239000003242 anti bacterial agent Substances 0.000 abstract description 12
- 239000000022 bacteriostatic agent Substances 0.000 abstract description 4
- 230000007613 environmental effect Effects 0.000 abstract description 3
- 239000002994 raw material Substances 0.000 abstract description 2
- 229940088710 antibiotic agent Drugs 0.000 description 7
- 244000052616 bacterial pathogen Species 0.000 description 4
- 230000002401 inhibitory effect Effects 0.000 description 4
- 239000000047 product Substances 0.000 description 4
- 241000894006 Bacteria Species 0.000 description 3
- 241000208368 Euonymus alatus Species 0.000 description 3
- 238000011161 development Methods 0.000 description 3
- 238000000034 method Methods 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- 206010059866 Drug resistance Diseases 0.000 description 2
- 241001465754 Metazoa Species 0.000 description 2
- 241000209140 Triticum Species 0.000 description 2
- 235000021307 Triticum Nutrition 0.000 description 2
- 230000003187 abdominal effect Effects 0.000 description 2
- 230000000975 bioactive effect Effects 0.000 description 2
- 235000013339 cereals Nutrition 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 230000035755 proliferation Effects 0.000 description 2
- 229930000044 secondary metabolite Natural products 0.000 description 2
- 241000606161 Chlamydia Species 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 241000222722 Leishmania <genus> Species 0.000 description 1
- 241000204031 Mycoplasma Species 0.000 description 1
- 241000422193 Ormyrus Species 0.000 description 1
- 241001478428 Syngnathus Species 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- 230000003042 antagnostic effect Effects 0.000 description 1
- 230000008485 antagonism Effects 0.000 description 1
- 230000000845 anti-microbial effect Effects 0.000 description 1
- 239000000427 antigen Substances 0.000 description 1
- 102000036639 antigens Human genes 0.000 description 1
- 108091007433 antigens Proteins 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000003115 biocidal effect Effects 0.000 description 1
- 210000002421 cell wall Anatomy 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 230000002559 cytogenic effect Effects 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 230000008260 defense mechanism Effects 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 229930183674 euplotin Natural products 0.000 description 1
- 230000009036 growth inhibition Effects 0.000 description 1
- 238000012423 maintenance Methods 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 239000002207 metabolite Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 244000052769 pathogen Species 0.000 description 1
- 230000001717 pathogenic effect Effects 0.000 description 1
- 230000000704 physical effect Effects 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 239000013535 sea water Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 230000002195 synergetic effect Effects 0.000 description 1
- 230000009897 systematic effect Effects 0.000 description 1
- 150000003505 terpenes Chemical class 0.000 description 1
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/66—Microorganisms or materials therefrom
- A61K35/68—Protozoa, e.g. flagella, amoebas, sporozoans, plasmodium or toxoplasma
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/04—Antibacterial agents
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- 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
- C12N1/10—Protozoa; Culture media therefor
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
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Abstract
The invention discloses a marine protozoan armillar extract with antibacterial activity, and relates to the technical field of protozoan extracts. The invention comprises a preparation method of the marine periclase extract and a micro dilution method for measuring the antibacterial activity of the marine periclase extract; the preparation method of the marine periwinkle extract is used for preparing the protozoan periwinkle extract; the application of determining the antibacterial activity of the marine periclase extract by a micro dilution method in preparing a bacteriostatic agent from the marine periclase antibacterial extract. The marine periclase extract has high antibacterial activity, can be used for preparing a bacteriostatic agent, is derived from marine protozoa, can continuously obtain raw materials through enlarged culture, and has environmental friendliness and low application cost; the antibacterial agent has a very broad application prospect and is used for developing antibacterial agents of methicillin-resistant staphylococcus aureus, vibrio rotifer, vibrio cansonii and vibrio vulnificus.
Description
Technical Field
The invention belongs to the technical field of protozoan extracts, and particularly relates to a marine protozoan euonymus alatus extract with antibacterial activity.
Background
Pathogenic microorganisms have great harm to human production and life, antibiotics are taken as secondary metabolites generated by microorganisms or higher animals and plants in the life process, and can kill bacteria and inhibit and kill other pathogenic microorganisms such as mould, mycoplasma, chlamydia and the like well, but in the past decades, the general use of antibiotics can cause pathogenic bacteria to generate drug resistance, and the development of novel antibiotics is more urgent due to the occurrence of antibiotic resistant bacteria;
over the past few decades, the availability of antibiotics has focused mainly on terrestrial resources, however the likelihood of finding new bioactive compounds from terrestrial organisms has been decreasing; marine environmental organisms are an important source of novel active secondary metabolites, and marine environments have been the main source of finding and finding new product resources because of their ecological pressure, including competition for space and predation, and physical properties such as salinity, pressure and temperature changes, which have led to the development of unique metabolic systems and organism defense mechanisms in this highly demanding ecosystem to adapt to such environments, whereby the characteristics of marine microorganisms must also be greatly different from those of land microorganisms, and thus marine organisms may produce a variety of new bioactive metabolites other than land;
the protozoa are eukaryotic unicellular animals without cell walls, and have all functions necessary for life maintenance and offspring extension although the structure is simple, and researches in the related fields are focused on aspects of morphology classification, systematics, cytogenetics, disease and ecology, and in recent years researchers have extracted some pathogen antagonistic active substances from some marine protozoa, such as the thick downstream servant of the order of the abdominal hairEuplotes crassus) Can produce a unique terpenoid named Euplotin, can resist the antagonism effect of Promastigotes on leishmania, and can be used as a synergistic of antigen worm chemotherapeutics in pharmacy. The study of the antimicrobial active ingredients of protozoa is of great importance.
Disclosure of Invention
Aiming at the problem that the pathogenic bacteria have drug resistance to the existing antibiotics due to the common use of the antibiotics, the invention provides the marine protozoan armillar extract with antibacterial activity, and provides resources for the research and development of novel antibiotics.
The invention is realized by the following technical scheme: an extract of marine protozoan Syngnathus with antibacterial activity comprises a preparation method of the extract and a micro dilution method for measuring antibacterial activity of the extract;
the preparation method of the marine periwinkle extract is used for preparing the protozoan periwinkle extract;
the micro dilution method is used for measuring the antibacterial activity of the marine periclase extract and detecting the antibacterial effect of the marine periclase extract.
Further, the preparation method of the marine periclase extract comprises the following steps:
the first step: separating and purifying the armillar insect population from the environment to obtain a single armillar insect population;
and a second step of: performing amplification culture on the bollworms, and inoculating a single species group of the bollworms into a liquid culture medium for amplification culture;
and a third step of: preparing extract, extracting with methanol, repeatedly extracting for 3 times, adding methanol into the sediment of the water sample removed after centrifugation, homogenizing the sediment by adopting a high-speed dispersion machine, centrifuging to be 12000r/min, taking a supernatant methanol sample, concentrating into extract by adopting a rotary evaporator, and then redissolving to be 5mL with methanol, and preserving at 4 ℃ for later use.
Further, the micro dilution method is used for measuring the antibacterial activity of the marine periwinkle extract, and the antibacterial activity of the marine periwinkle extract is measured by detecting the inhibition percentage of thalli added into the marine periwinkle extract in the culture of pathogenic microorganisms.
Further, the microorganism is methicillin-resistant staphylococcus aureus, vibrio rotifer, vibrio cansonii and vibrio vulnificus.
The invention has the following beneficial effects:
the marine periclase extract has high antibacterial activity, can be used for preparing a bacteriostatic agent, is derived from marine protozoa, can continuously obtain raw materials through enlarged culture, and has environmental friendliness and low application cost; the antibacterial agent has a very broad application prospect and is used for developing antibacterial agents of methicillin-resistant staphylococcus aureus, vibrio rotifer, vibrio cansonii and vibrio vulnificus.
Detailed Description
The following description of the technical solutions in the embodiments of the present invention will be clear and complete, and it is obvious that the described embodiments are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
The invention relates to a marine protozoan armillar extract with antibacterial activity, which comprises a preparation method of the marine armillar extract and a micro dilution method for measuring the antibacterial activity of the marine armillar extract;
the preparation method of the marine periwinkle extract is used for preparing the protozoan periwinkle extract;
the preparation method of the marine periclase extract comprises the following steps:
the first step: separating and purifying the euonymus alatus population from the environment;
collecting protozoa community from seawater, expanding the community by using wheat grain culture solution in a laboratory, and the operation steps are as follows: sucking a bollworm under microscopeGastrostyla sp.), performing expanded culture by using wheat grain culture solution to obtain a single species group of the armillar worms;
and a second step of: expanding culture of the euonymus alatus;
inoculating single species of Bulleyan in liquid culture medium, shake culturing at 25deg.C for 2 months at low speed, and performing amplification culture to 5L scale, centrifuging (3,000 rpm,10 min) the obtained sample, collecting insect body, inoculating single species of Bulleyan in liquid culture medium, and performing amplification culture;
and a third step of: preparing an extract; extracting with methanol for 3 times repeatedly, adding methanol into the sediment of the water sample removed after centrifugation, homogenizing the sediment by adopting a high-speed dispersing machine, centrifuging to 12000r/min, collecting a supernatant methanol sample, concentrating into an extract by adopting a rotary evaporator, re-dissolving to 5mL by using methanol, and preserving at 4 ℃ for later use;
high speed disperser was purchased from the product number of the laboratory technology, guangzhou department of technology, IKA Ai Ka, germany: t25DS25;
rotary evaporator was purchased from Heidolph, germany, product number: hei-VAP;
the application of the micro dilution method for measuring the antibacterial activity of the marine periclase extract in preparing antibacterial agent by using the marine periclase antibacterial extract.
The method for determining the antibacterial activity of the marine periwinkle extract by using the micro dilution method adopts the method for detecting the growth inhibition condition of the thalli added with the marine periwinkle extract in the culture of pathogenic microorganisms to determine the antibacterial activity of the marine periwinkle extract.
The microorganism is methicillin-resistant staphylococcus aureus, vibrio rotifer, vibrio candidum and vibrio vulnificus;
two strains of methicillin-resistant staphylococcus aureus, vibrio rotifer, vibrio cankeri and vibrio vulnificus. By OD 600 Determining the minimum inhibitory concentration MIC at 90% inhibition of each species;
the above strains are available from ATCC, CGMCC and MCC;
the minimum inhibitory concentration MIC of the marine periclase extract on the pathogenic bacteria is shown in table 1, and from table 1, the marine periclase extract effectively inhibits the proliferation of the pathogenic bacteria, has higher activity of inhibiting the proliferation of bacteria, can be used for preparing a bacteriostatic agent, and has very broad development and application prospects;
TABLE 1 MIC results Table of minimum inhibitory concentration of Marine Abdominal column extract
The reagents or apparatus used were conventional products commercially available without the manufacturer's attention.
In the description of the present specification, the descriptions of the terms "one embodiment," "example," "specific example," and the like, mean that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the present invention. In this specification, schematic representations of the above terms do not necessarily refer to the same embodiments or examples. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples.
The preferred embodiments of the invention disclosed above are intended only to assist in the explanation of the invention. The preferred embodiments are not exhaustive or to limit the invention to the precise form disclosed. Obviously, many modifications and variations are possible in light of the above teaching. The embodiments were chosen and described in order to best explain the principles of the invention and the practical application, to thereby enable others skilled in the art to best understand and utilize the invention. The invention is limited only by the claims and the full scope and equivalents thereof.
Claims (1)
1. An extract of marine protozoan armillar with antibacterial activity, which is characterized in that: the preparation method comprises the steps of preparing the marine periclase extract, and determining the antibacterial activity of the marine periclase extract by a micro dilution method after the preparation is finished;
the preparation method of the marine periwinkle extract is used for preparing the protozoan periwinkle extract;
the micro dilution method is used for measuring the antibacterial activity of the marine periclase extract;
the preparation method of the marine periclase extract comprises the following steps:
the first step: separating and purifying the armillar insect population from the environment to obtain a single armillar insect population;
and a second step of: performing amplification culture on the bollworms, and inoculating a single species group of the bollworms into a liquid culture medium for amplification culture;
and a third step of: preparing an extract, extracting with methanol for 3 times repeatedly, wherein the extraction method comprises adding methanol into the sediment of a water sample removed after centrifugation, homogenizing the sediment by a high-speed dispersion machine, centrifuging to 12000r/min, collecting a supernatant methanol sample, concentrating into an extract by a rotary evaporator, and then redissolving to 5mL with methanol, and preserving at 4 ℃ for later use;
after the preparation is finished, determining the antibacterial activity of the marine periclase extract by the micro dilution method, and determining the antibacterial activity of the marine periclase extract by detecting the inhibition percentage of thalli added with the marine periclase extract in the cultured pathogenic microorganisms;
the microorganism is methicillin-resistant staphylococcus aureus, vibrio rotifer, vibrio candidum and vibrio vulnificus.
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| CN202111262517.4A CN113813292B (en) | 2021-10-28 | 2021-10-28 | Marine protozoan armillar extract with antibacterial activity |
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1997034489A1 (en) * | 1996-03-20 | 1997-09-25 | Aquaculture Technology, Inc. | Antibacterially active marine algae extracts from the class diatomatae and methods of use |
| JP2010059100A (en) * | 2008-09-04 | 2010-03-18 | Yokohama City Univ | Antibacterial agent for gram-positive bacteria and antimicrobial activity potentiating agent |
| KR20120063245A (en) * | 2010-12-07 | 2012-06-15 | 전남대학교산학협력단 | Anti-bacterial and anti-protozoal composition containing bromophenol derivatives |
| CN108283653A (en) * | 2018-04-05 | 2018-07-17 | 吉林大学 | It is a kind of using Taxus extracts prepare plant-based bacteriostat method and its in the application of antibiosis |
| CN109576341A (en) * | 2018-12-05 | 2019-04-05 | 海南省海洋与渔业科学院（海南省海洋开发规划设计研究院） | A kind of lead compound screening technique inhibiting grouper pathogenic vibrio |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| US11851644B2 (en) * | 2019-05-02 | 2023-12-26 | University Of Rhode Island Board Of Trustees | Marine bacteria formulation useful in aquaculture |
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2021
- 2021-10-28 CN CN202111262517.4A patent/CN113813292B/en active Active
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| CN109576341A (en) * | 2018-12-05 | 2019-04-05 | 海南省海洋与渔业科学院（海南省海洋开发规划设计研究院） | A kind of lead compound screening technique inhibiting grouper pathogenic vibrio |
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