159 results about "Tetrodotoxin" patented technology

This invention relates to a stable pharmaceutical composition of freeze-dried tetrodotoxin powder which contains trace amount of tetrodotoxin, substances which can stabilizes tetrodotoxin, including disaccharide(s) or polyglucose(s) or analogues thereof and solvent(s), and solvents which can help tetrodotoxin dissolve.

A medicine in the form of aerosol or spray for dropping drug or antalgic purpose is prepared from tetrodotoxic citrate or tetrodonic acetate. Its advantages are applying it via respiratory tract, high effect up to 97% or more, high safety, and no addiction.

The present invention relates to matter extraction from animal material and is the extraction process of tetrodotoxin. The present invention provides the technological method of raising yield of tetrodotoxin. The technological process includes the steps of: cutting internal organs of globefish, grinding into slurry, press filtering, heating to eliminate protein and oil, filtering to obtain clear liquid, concentration, elution of crystal inchromatographic column to obtain pure tetrodotoxin product.

The invention relates to a method for separating Aeromonas molluscorum producing tetrodotoxin from Takifugu fasciatus tissues and a fermentation culture method of the Aeromonas molluscorum as well as a detection method of the produced tetrodotoxin. The method for separating the Aeromonas molluscorum producing the tetrodotoxin from the Takifugu fasciatus tissues comprises a bacteria separation method, a bacteria identification method, a bacteria fermentation culture method, a competitiveness ELISA (Enzyme Linked Immunosorbent Assay) detection method and an LC-MS (Liquid Chromatograph-mass Spectrometry) detection method of the tetrodotoxin produced by the Aeromonas molluscorum, and the like. In the invention, by separating bacteria from Takifugu fasciatus ovaries and adopting a TCBS (Thiosulfate Citrate Bile (salt) Sucrose (agar)) culture medium for screening, the Aeromonas molluscorum capable of producing the tetrodotoxin is separated from blowfish bodies for the first time; the separated bacteria is identified by adopting a 16 SrDNA method; the competitiveness ELISA detection method and the LC-MS detection method are both firstly adopted for the tetrodotoxin produced by the separated Aeromonas molluscorum; and the produced tetrodotoxin and tetrodotoxin extracted from the blowfish bodies are determined to be the same substance; in addition, the separated Aeromonas molluscorum can massively separate the tetrodotoxin from the bacteria after being cultured.

A process for preparing the liquid extract of tetrodotoxin includes such steps as quickly freezing the viscera of globefish, cutting to become small blocks, immersing in deionized water, fermenting, quick heating, quick cooling, primary filtering and fine filtering. It has high output rate.

The externally applied compound tetrodotoxin ointment is prepared with globe fish liver oil, vaseline, frankincense, myrrh, dragon's blood, notoginseng, safflower, catechu, borneol, vitamin B1, vitamin B2, vitamin C, moroxydine, dexamethasone and lidocaine in certain weight proportion. It has very high curative rate on trauma and traumatic infection, including intractable ulcer and fistula, high effective rate and curative rate on skin diseases caused by bacteria, fungi, viruses, etc, and certain treating effect on serious furuncle, luetic balanitis, etc.

The invention discloses a tetrodotoxin quick-release pellet preparation and its preparation method and use and relates to the field of a medicinal preparation technology and use. The tetrodotoxin quick-release pellet preparation is suitable for oral administration and utilizes a blank pellet core as a carrier and a tetrodotoxin aqueous solution containing a cosolvent and a binder as a drug feeding solution. A weight ratio of the tetrodotoxin to the pellet core is 0.0025%-0.3% and an oral dosage of the tetrodotoxin is less than or equal to 300 micrograms. The tetrodotoxin quick-release pellet preparation has the advantages of high superimpose rate, good content uniformity, fast drug release rate and fast pain easing. The tetrodotoxin quick-release pellet preparation has the characteristics of good clinical use compliance and high safety. The blank pellet core fluidized bed-based drug superimpose method is suitable for preparation of a very low specification tetrodotoxin oral preparation.

The invention discloses a process for preparing leftover fertilizer processed through globefish by adopting an enzymolysis and fermentation method. A composite enzymatic hydrolysis and multi-strain fermenting mode is adopted for degrading macromolecule substances difficult to utilize in leftovers processed through the globefish into micromolecule polysaccharide, polypeptide and nitrogen-containing compounds, a large number of soil beneficial bacteria and various special-effect metabolites are generated, the biological activity of the leftover fertilizer processed through the globefish is improved, and pesticide and the fertilizer are combined into one, which is beneficial for pushing the development of the ecological agriculture and increasing the resource comprehensive utilization rate and the economic benefits of the globefish processing industry. The number of the effective active bacteria in the fertilizer is larger than or equal to 20 billion / mL. The fertilizer integrates natural nutrient ingredients, antibiotic substances and zoohormone, and has a composite effect of organic fertilizer, microbial fertilizer and microelement fertilizer; the fertilizer contains a certain amount of tetrodotoxin, and can effectively kill pests in soil, the pesticide and fertilizer combination is achieved, the duration time for the pest killing effect is long, and the application prospect is wide.

The invention discloses a method for determining tetrodotoxin in marine organisms by utilizing immunoaffinity column purification-liquid phase chromatography-tandem mass spectrometry. The method is characterized in that the tetrodotoxin in the marine organisms can be rapidly, simply and high efficiently determined by utilizing the unique selection identifiability of the immunoaffinity column on the basis of the high sensitivity and accuracy of the liquid phase chromatograph tandem mass spectrometry.

The invention provides a set of single stranded DNA (ssDNA) aptamers DA-01 and TTX-27 which can simultaneously recognize domoic acid (DA) and tetrodotoxin (TTX), including one ssDNA aptamer DA-06 which specifically recognizes DA, one ssDNA aptamer TTX-07 which can specifically recognize TTX, and one ssDNA aptamer STX-41 which can specifically recognize saxitoxin (STX). According to the invention,magnetic reduced graphene oxide (MRGO) is adopted to assist in separated systematic evolution of ligands by exponential enrichment (SELEX), properties of MRGO of being capable of adsorbing free ssDNA,but not adsorbing ssDNA combined with target molecules are utilized, and magnetic separation enrichment characteristics of MRGO are further used to separate ssDNA with affinity and without affinity to target toxins; primary structure homology and secondary structure similarity on obtained ssDNA are analyzed after 16 repeated rounds of incubation, separation, amplification and in-vitro screening for single-strand preparation, then the affinity and the specificity are measured, and finally one set of the aptamers with high affinity and specificity is obtained. The set of the aptamers has broadapplication prospects in the aspects of analysis, detection, separation, enrichment, removal and purification for DA, TTX and STX in aquatic products.

The invention discloses a rapid cultivation feed for takifugu rubripes. The rapid cultivation feed comprises the following components in parts by weight: 20-40 parts of domestic fish meal, 10-20 parts of shrimp powder, 10-20 parts of meat and bone meal, 1-10 parts of chicken oil, 10-20 parts of corn starch, 1-5 parts of soybean lecithin, 0.1-5 parts of an oyster shell extract, 0.1-8 parts of cylindrotheca fusiformis powder, 2-6 parts of sodium alginate, 1-15 parts of functional additive components, 0.1-3 parts of multivitamins and 0.1-5 parts of multiminerals. A preparation method of the rapid cultivation feed disclosed by the invention comprises the following steps: (1) manufacturing raw materials; (2) synthesizing a biological material; (3) converting to a cooked material; (4) puffing and granulating; (5) drying; (6) spraying and coating; and (7) performing vacuum drying. According to the feed disclosed by the invention, by scientific matching, the body color of adult takifugu rubripes can be improved, the growth can be promoted, and the enrichment of tetrodotoxin can be reduced.

The domesticating and culturing process of dark-grained Eastern globefish with controlled tetrodotoxin includes the steps of artificial propagation, fry culture, globefish culture, etc. The present invention makes it possible to control the tetrodotoxin content under fresh water culture condition simply without pollution.

The invention provides a method for producing tetrodotoxin from microorganisms by commensalism culture fermentation. The method is characterized by comprising the following steps: preparing microorganism strains which are separated from veneniferous tissues of a wild puffer into bacterial suspensions by using sterile distilled water, and mixing the bacterial suspensions and inoculating into a same liquid culture medium to carry out seed liquid culture for 1-3 days; and inoculating 5-10% inoculation amount of seed liquid into a fermentation culture medium to carry out fermentation culture for 2-7 days, wherein the liquid culture medium comprises the following components by weight percent: 0.1-0.5% of yeast extract, 0.1-0.5% of beef extract, 0.1-0.5% of peptone, 0.5-2% of glucose, 0-1% of sugar and the balance of seawater, and the pH value is 6.5-7.2; and the seed liquid is subjected to fermentation culture under the conditions that the temperature is 24-30 DEG C and the rotation speed is 150-200 rpm (revolutions per minutes). According to the method provided by the invention, a micro-ecological environment in which the microorganisms producing toxicity in the puffer is located can be well simulated, thereby being beneficial to the adaptability of the microorganisms producing toxicity to culture in vitro, finally being beneficial to growth and toxicity production of the microorganisms producing toxicity and solving the bottleneck that at present, the yield of the tetrodotoxin produced by microbial fermentation is low in general.

The invention relates to a method for preparing a tetrodotoxin-degrading substance from a tetrodotoxin-producing bacteria fermentation broth, and a preparation and detection method of an extract liquid. The method comprises a method for artificially causing plasmid loss, a tetrodotoxin-degrading substance extraction method, and a tetrodotoxin degradation detection method. According to the invention, for a first time, the existence of a tetrodotoxin-degrading substance in aeromonas molluscorum is found, and an extraction method of the substance is established. The method has certain potential to be used in large-scale productions of the tetrodotoxin-degrading substance. The extract of the bacteria fermentation broth has a tetrodotoxin-degrading effect, and can be used for preparing medicines used for treating tetrodotoxin intoxication. Also, a raw material can be provided for related fields such as scientific research and pharmaceutical industries.

The invention discloses a formula feed for juvenile fish of takifugu rubripes and a preparation method of the formula feed. The feed contains the following components in parts by weight: 30-50 parts of whitefish meal, 10-20 parts of krill meal, 10-20 parts of earthworm powder, 1-10 parts of squid oil, 1-10 parts of hordein powder, 1-10 parts of rice bran, 2-12 parts of dried yellow wine lees, 5-15 parts of wheat germ oil, 1-5 parts of evening primrose seed oil, 0.1-0.5 part of soybean enzymolysis protein, 0.1-0.5 part of a mussel extract, 1-8 parts of schizochytrium limacinum powder, 2-6 parts of nannochloropsis oculata powder, 5-15 parts of functional additive components, 0.1-0.3 part of multivitamins and 0.1-0.5 part of multiminerals. The preparation method comprises the following steps: (1) manufacturing raw materials; (2) synthesizing a biological material; (3) converting to a cooked material; (4) puffing and granulating; (5) drying; (6) spraying and coating; and (7) performing vacuum drying. The feed disclosed by the invention can improve the survival rate of the juvenile fish of takifugu rubripes, improve the physique and the body color, promote the growth, reduce the enrichment of tetrodotoxin and reduce the cultivation cost.

The invention relates to a tetrodotoxin preparation for treating drug addiction and relieving pain. The preparation comprises small-dose tetrodotoxin serving as a medicinal active ingredient and a pharmaceutical permissible cosolvent or support agent, wherein the cosolvent is amino acid. The amino acid is used as the cosolvent, so the amino acid has the function of the cosolvent, increases the water solubility of the tetrodotoxin, and can also form highly water-soluble salt solution with the tetrodotoxin. The tetrodotoxin used in the invention is prepared by a specific method; and the purity of the prepared tetrodotoxin reaches 98 percent and the yield is high. The preparation consisting of the tetrodotoxin prepared by the method and the amino acid has the advantages of high water solubility, high stability, safety, reliability, stable quality, no need of preservation in a refrigerator at a specific temperature, storage life of more than two years at room temperature, capability of greatly saving the storage and transportation cost and facilitating medical use, obvious treatment effect, and cure rate of 100 percent when particularly used for physiological detoxification.

The invention discloses a preparing method for tetrodotoxin standard biological samples. The preparing method includes the steps that nassarius samples are extracted with an acetic-acid methanol solution, a crude extracting solution is purified with a carbon-nano-tube solid-phase extraction column, the purified venom is subjected to high-performance-liquid-chromatography purification, and then isfrozen, dried and concentrated, crystals are separated out, and the tetrodotoxin standard biological samples are obtained. According to the preparing method for the tetrodotoxin standard biological samples, in the TTX extraction aspect, the purifying aspect and the pure-product preparing aspect, the nassarius serves as a raw material, extraction is carried out with the acetic-acid methanol solution, the crude extracting solution is purified through the large-volume-carbon-nano-tube solid-phase extraction column, then the purified venom is concentrated, and the samples are purified and preparedthrough high-performance-liquid sample feeding. The sample preparing method and process are easy and convenient to operate, efficient and suitable for batch production and technology preparation of the tetrodotoxin standard biological samples.

The invention discloses microdiet for fish larvae of takifugu rubripes. The microdiet comprises components as follows: white fish meal, tenebrio molitor meal, earthworm powder, shrimp oil, rice protein, husked sorghum flour, lotus root starch, maize germ oil, walnut oil, casein hydrolysate, green-lipped mussel extract, cylindrotheca powder, haematococcus pluvialis powder, a functional additive, complex vitamins and compound minerals. The preparation method of the microdiet comprises steps as follows: 1), preparation of raw materials; 2), synthesis of biological materials, 3), conversion into cooked materials; 4), puffing for granulation; 5), drying; 6), spraying for film coating; 7), vacuum drying. The microdiet can improve the body color of the fish larvae of the takifugu rubripes, promote growth and reduce tetrodotoxin enrichment.

The invention provides an expression vector and a preparation method thereof, and high-purity recombinant HNTX (Hainantoxin)-IV analogue rHNIV-01 with biological activity can be rapidly prepared by the expression vector. Compared with natural HNTX-IV, an amino acid sequence of rHNIV-01 is almost the same as that of natural HNTX-IV, and only the C terminal of rHNIV-01 is not subjected to amidation modification. The method does not adopt conventional affinity chromatography for purifying fusion protein, protein kinase is directly added in a cell lysis product for cutting, then a cutting product rHNIV-01 is directly subjected to selective active extraction with a TCA (trichloroacetic acid) extraction method, and finally, RP-HPLC (reversed phase high-performance liquid chromatography) is adopted for purification. With the adoption of the method, 3 mg / L high-purity rHNIV-01 can be obtained stably; and purified rHNIV-01 can selectively inhibit TTX-S (tetrodotoxin-sensitive) sodium current on DRG (dorsal root ganglia) cells of a rat.

The invention discloses a making method of a semi-quantitative tetrodotoxin detection card. The method comprises the following steps: 1, preparing a colloidal gold sample combination pad; 2, chromatographic membrane processing: uniformly drawing a tetrodotoxin artificial antigen forming a detection line and a quality control antibody forming a quality control line in the middle of a chromatographic membrane by using a membrane drawing instrument; and 3, assembling: splicing a carrier pad, the chromatographic membrane, the colloidal gold sample combination pad and a water absorption pad to form a layered structure, wherein the carrier pad is positioned at the bottom, the chromatographic membrane is arranged on the carrier pad, and the colloidal gold sample combination pad and the water absorption pad are respectively placed at two ends of the chromatographic membrane. The above obtained product allows tetrodotoxin to be rapidly semi-quantitatively analyzed through ocular estimation, and has the characteristics of small size, light weight, easy preservation and transportation, and realization of detection of a detected sample without separation or purification.

The invention relates to a method for extracting tetrodotoxin from globefish internal organ tissues and then primarily purifying the tetrodotoxin. The globefish internal organ tissues are subjected to homogenate, dialysis, and reversed-phase weak anionic chromatography to remove the impurities, then tetrodotoxin is condensed and purified by a weak cationic exchange technology, and finally the tetrodotoxin is subjected to freeze drying so as to obtain the tetrodotoxin dry powder with a purity of 80%. In the provided method and technology, LC-MS or LC-MS / MS is used to detect the tetrodotoxin concentration in each step so as to guarantee that no tetrodotoxin is lost or discarded. The tetrodotoxin yield of the provided method is 10 to 100 times more than that of conventional methods recorded in literature and current patents. The provided technology especially the continuous fluid and special weak cationic exchange materials can process 10 kilograms of globefish ovary at one time, and is capable of obtaining 10 to 20 grams of tetrodotoxin dry powder with a purity of 80% to 90%.

The present invention refers to the use of a sodium channel blocker such as tetrodotoxin or saxitoxin, their analogues and derivatives as well as their physiologically acceptable salts, in medicinal products for human therapeutics for the treatment of central-nervously derived neuropathic pain.

The invention provides methods for treating visceral pain and pain associated with therapy. The compounds useful in the methods of the invention are blockers of sodium ion channels, and in particular compounds that bind to the SS1 or SS2 extracellular mouth of the α-subunit thereof. Particularly useful compounds are saxitoxin and its derivatives and analogues and tetrodotoxin and its derivatives and analogues.

The method for extracting tetrodotoxin from liver of globefish includes the following steps: cutting liver of globefish, soaking at low temp., stirring, filtering, freezing, repeatedly soaking for several times to obtain crude venom, freezing and standing still, precipitating, heating and boiling, discharging out venom, freezing, and separating out supernatant venom, regulating pH value, adding active carbon, stirring, filtering, mixing cation 110 resin and said clear liquor, stirring, filtering, making said clear liquor pass through cation 110 resin, saturating and using acid to make elution, collecting eluant, regulating pH value, cooling, standing still, crystallizing, sucking out mother liquor, filtering, more regulating pH, freezing, standing still, crystallizing and repeatedly regulating pH for several times to obtain white powder crystal.

The present invention relates to a tetrodotoxin preparation used for drugs abstinence and analgesia. The preparation comprises small dose of tetrodotoxin that is taken as active ingredient of medicine and cosolvent or allowable supportting agent in pharmacy. The cosolvent is amino acid. In the present invention, the amino acid is taken as the cosolvent, which not only can act as the cosolvent for increasing solubility of the tetrodotoxin in water, but also can form highly water-soluble salting liquid with the tetrodotoxin. The tetrodotoxin used in the present invention is prepared by adopting the specific method of the present invention. The purity of the prepared tetrodotoxin is as high as 98 percent with high yield. The preparation formed by the tetrodotoxin prepared by adopting the method provided in the present invention and amino acid is not only good in water solubility, but also good in stability, safe, reliable, stable in quality. The preparation does not need to be preserved in the refrigerator at specific temperature. The preparation can be preserved for more than 2 years at room temperature, thus greatly saving storage and transportation cost and providing convenience for use in medical treatment. The cure effect is evident, and in particular when applied to physiologic de-toxin, the tetrodotoxin preparation has a cure rate reaching 100 percent.

The invention relates to a method for massively and extremely efficiently extracting high-purity tetrodotoxin from viscera of globefishes, and aims at overcoming defects, in existing various methods for extracting and purifying tetrodotoxin, that the efficient liquid phase method with high product purity is high in cost of equipment (a high-pressure liquid phase instrument), and the dipping fractionation method is low in efficiency and low in product purity. The tetrodotoxin is applicable to drug rehabilitation, analgesia and pressure reduction drug preparations. The method is performed for massively extracting and preparing tetrodotoxin. The method is simple in processes, high in efficiency, high in yield, and high in quality; the product purity is up to 97-99% and directly meets the requirements of biological study and clinical application. Additionally, the invention aims at providing various preparations containing the tetrodotoxin prepared by the method.

The present invention relates methods for the biosynthesis of tetrodotoxin (TTX) involving the steps of obtaining a culture possessing one or more of a Vibrio species, such as through a seed culture / inoculating the culture and tissue extract from a textrodotoxin-bearing organism in a fermenter medium, and isolating and purifying tetrodotoxin from said fermenter, resulting in a yield of TTX of about 0.5 g TTX / L in about 3 to 5 days, such TTX being at least 90% pure.

The invention discloses a method for rapidly detecting the content of tetrodotoxin in fish flesh through liquid chromatography-mass spectrometry. The method comprises (1) sample preprocessing and (2) chromatography-mass spectrometry detection. Diatomite is added in the preprocessing process against the effect problem of a matrix in a fish flesh sample and combining the chemical characteristics of tetrodotoxin, and the tetrodotoxin is separated from muscle tissues in an ultralow temperature freezing manner. A sample undergoes extraction by 1% acetic acid-methanol in a cell crasher at 60 DEG C, obtained extract is purified by a Carbon-GCB SPE small column, and the purified extract is directly introduced and analyzed, so the concentration time is saved. Compared with 0.2% acetic acid-water solution extraction, other SPE purification and ultrafiltration methods in the prior art, the method disclosed in the invention, adopting direct introduction and analysis of the sample, has the advantages of simplicity and low consumption in the processing process, good accuracy, convenience in operation, and good reappearance.

The invention provides a tetrodotoxin enteric-coated and sustained-release pellet and a preparation method thereof, and relates to the field of a medicine preparation technology and application. The tetrodotoxin enteric-coated and sustained-release pellet sequentially comprises a blank pellet core, an upper medicine layer, a sustained-release layer, an isolation layer and an enteric layer from inside to outside, wherein the upper medicine layer is tetrodotoxin containing a cosolvent and a bonding agent; according to a weight ratio, the weight of the sustained-release layer accounts for 20 percent to 40 percent of the weight of the medicine containing pellet; the weight of the isolation layer accounts for 2 percent to 8 percent of the weight of the sustained-release pellet; the weight of the enteric layer accounts for 10 percent to 30 percent of the weight of the isolation pellet. The blank pellet core is used as a medicine carrier; a fluidized bed medication method is used for preparing the tetrodotoxin medicine-carried pellet; the sustained-release layer, the isolation layer and the enteric layer are sequentially sprayed to obtain the tetrodotoxin enteric-coated and sustained-release pellet. The advantages of medicine release controllability, a long-period pain relieving effect, good clinic use complaisance, high safety and the like are realized. In addition, the blank pellet core fluidized bed medication and coating method is applicable to the preparation of the ultra-low-specification tetrodotoxin medicine oral sustained-release preparation; the medication rate is high; the content uniformity is good.

The present invention refers to the use of sodium channel blockers such as tetrodotoxin or saxitoxin, its analogues / derivatives as well as their acceptable salts, for the production of a medicament for the treatment of neuropathic pain resulting from chemotherapy.