Method of obtaining natural 9-decen-2-one by bioconversion of undecylenic acid by means of mould and application in perfumery as food fragrance
SUBSTANCE: invention relates to method of obtaining 9-decen-2-one. Method includes cultivation of said mould, belonging to family Aspergillaceae or Mortierellaceae, addition of undecylenic acid as substrate in fermentation medium with consumption from 0.1 to 0.9 g/l/hour in presence of oil, substrate bioconversion in 9-decen-2-one, its extraction and purification. As oil, used is oil, selected from the group, containing traditional food oils or triglycerides, which contain short-chained fatty acids, or white oils.
EFFECT: invention makes it possible to obtain 9-decen-2-one with output 8-16,5 g/l and high degree of purification 99%.
13 cl, 1 dwg, 9 ex
The present invention relates to the bioconversion using a mold undecylenate acid 9-mission-2-it, the latter can be used in perfumery and as a food flavoring.
The applicant found that 9-mission-2-he (or decene-2 or methylacetanilide, CAS RN 35194-30-0) is a ketone having such olfactive and gustatory properties that it can be used for all applications in the production of flavors (flavors and odors). Indeed he has a green oily note, and fruit notes, particularly pear, Apple, pineapple, passion fruit exotic fruit in General.
This ketone as well as other methylketone, formed pathway, which involves the first stage of β-oxidation after oxidation of fatty acids. This metabolic pathway is the formation of methylketones used mushrooms in detoxification of fatty acids.
As you can see from the image metabolic pathway (Fig.1), the substrate used in the process of bioconversion is undecylenate acid, which is produced by cracking (physical operation of transforming crude product in a more purified compounds) ricinoleic acid derived from castor oil, followed by purification by dry distillation. Finally, after phase decarboxylation of matrceta is, received in the end of the path contains one carbon atom less than the original fatty acid.
As far as known to the applicant, to date there is no biotechnological process that describes and/or allows the synthesis of 9-mission-2-it. Indeed, to date, this molecule can only be obtained by chemical synthesis and minor release. In addition, the lack of synthetic fragrances is that they are less valued by consumers than the natural aromas. Therefore, the aim of the present invention is an alternative method of obtaining natural aromatic molecules, in particular, obtained using 9-mission-2-it, by biological means, more specifically, by bioconversion with the use of microorganisms such as mold.
In the context of the present invention, "bioconversion" understand the biological transformation of the substrate, preferably derived from a natural source, for the natural aromatic substances that can be used in recipes as fragrances and odors, which are natural.
Unexpectedly, the applicant found that this objective can be achieved by way of getting 9-mission-2-it is characterized by bioconversion undecylenate acid using p is esani, moreover, this method includes the following stages:
a) culturing a specified mold;
b) adding undecylenate acid as substrate bioconversion in the presence of oil;
C) bioconversion of the substrate in the 9-mission-2-it,
d) extraction and purification 9-mission-2-it.
In the context of the present invention, the term "mold" refers to microscopic filamentary single or multicellular fungi. Of the types of mold there are phykomycets and ceptability. As part of phykomycets there is a group of oomycetes and group zygomycetes. As part of satemization allocate a group of basidiomycetes and group deuteromycetes.
In the embodiment of the invention the mold according to the invention belongs to the group of deuteromycetes, class hyphomycetes, order Tuberculariales, family Aspergillaceae. More preferably the mould belongs to the genus Aspergillus or Penicillium. In the first preferred embodiment, the mould belongs to the species Aspergillus oryzae or to the species Aspergillus nyger. In the second preferred embodiment, the mould belongs to the species Penicillium roquefortii or to the species Penicillium camenbertii.
From the mold Aspergillus oryzae include collectible strains: Aspergillus oryzae DSMZ 1861, Aspergillus oryzae DSMZ 1864, Aspergillus oryzae DSMZ 1147, Aspergillus oryzae DSMZ 63303, Aspergillus oryzae CBS 570.65, Aspergillus oryzae CBS 819.72, Aspergillus oryzae CBS 110.27.
From the mold Aspergillus nyer include collectible strains: Aspergillus nyger DSMZ 823, Aspergillus nyger DSMZ 2466.
From the mold Penicillium roquefortii include collectible strains: Penicillium roquefortii CBS 221-30, Penicillium roquefortii PRB 18, Penicillium roquefortii DSMZ 1079, Penicillium roquefortii DSMZ 1080.
From the mold Penicillium camembertii can be called the strain Penicillium camembertii DSMZ 1233.
In another preferred embodiment, the mold according to the invention belongs to the group of zygomycetes, class Zygomyctes, order Mucorales, family Mortierellaceae or family tuberculariaceae, genus Fusarium.
More preferably the mould belongs to the genus Mortierella or the genus Mucor, the genus Rhizopus, and even more preferably the mold refers to a type of Mortierella isabellina or Mortierella rammniana, or to the species Mucor racemosus or Mucor miehei, or to the species Rhizopus arrihizus or Rhisopus oryzae.
From a fungus belonging to the species Mortierella, you can call the following collection strains: Mortierella isabellina DSMZ 1414, Mortierella isabellina CBS 100559, Mortierella isabellina CBS 221.29, Mortierella isabellina CBS 194.28, Mortierella isabellina CBS 208.32, Mortierella isabellina CBS 224.35, Mortierella isabellina CBS 560.63, Mortierella isabellina CBS 167.80, Mortierella isabellina CBS 493.83, Mortierella isabellina CBS 309.93, Mortierella isabellina CBS 250.95, Mortierella isabellina CBS 109075, Mortierella ramanniana CBS 112.08, Mortierella ramanniana CBS 219.47, Mortierella ramanniana CBS 243.58, Mortierella ramanniana CBS 478.63, Mortierella ramanniana CBS 852.72, Mortierella ramanniana CBS 366.95, Mortierella ramanniana CBS 101226.
From a fungus belonging to the species Mucor, you can call the following collection strains: Mucor javanicus DSMZ 1222, Mucor racemosus DSMZ 62760, Mucor rouxii DSMZ 1191.
From a fungus belonging to the species Rhizopus, you can call clubusacasino strains: Rhizopus arrhizus oryzae DSMZ 905, Rhizopus oryzae DSMZ 2199, Rhizopus deltmar DSMZ 853, Rhizopus niveus DSMZ 2194, Rhizomucor miehei DSMZ 1330.
Cultivation at the stage a) of the method according to the invention includes the creation of culture, preferably policecontributing, for example, by cell amplification in the respective cultural environment. This culture is preceded by a pre-culture of the strain in the first culture medium, more adapted to the first stages of reproduction.
In a preferred embodiment of the invention the residence time in the culture in this mold is from 20 to 30 hours, and in fact depends on the state of growth of the mycelium, and the latter can be observed in the microscope. Preferably the minimum biomass of the mold ranges from 7 to 15 g/l and more preferably from 9 to 12 g/l prior to the commencement of bioconversion according to the invention.
Stage b) of the method consists of adding a substrate for cell culture. According to the invention for the biological synthesis of 9-mission-2-it uses the most appropriate substrate, which is undecylenoyl acid or one of its esters, methyl ester or ethyl ester undecylenate acid. Needless to say, the substrate may be a mixture of a variety of suitable substrates, in particular, a mixture undecylenate acid and one or more of its esters.
In accordance with the advantages to the public of the embodiment of the invention, the substrate is introduced into the mycelium periodic manner or method injection.
In a preferred embodiment of the invention specified undecylenoyl acid is introduced into the fermentation medium with a flow rate of from 0.1 to 0.9 g/l/h, preferably from 0.2 to 0.7 g/l/h and very preferably from 0.3 to 0.55 g/l/h.
Stage C) of the method consists in the bioconversion undecylenate acid 9-mission-2-it. The duration of this stage of bioconversion mainly ranges from 36 to 96 hours.
Due to the high toxicity undecylenate acid for Mizerov the adjustment period may be necessary to prepare a mold for conversion of the substrate. Thus, in accordance with the preferred embodiment stage bioconversion method according to the invention includes the first phase of adaptation of the mould, during which the specified undecylenoyl acid is injected with a flow rate of from 0.1 to 0.5 g/l/h, then the second phase, during which undecylenoyl acid is injected with a flow rate of from 0.25 to 0.9 g/l/h. Mainly the duration of this stage adaptation of less than 20 hours, preferably less than 12 hours and more preferably less than 6 hours.
In accordance with the preferred embodiment undecylenoyl acid is administered in the presence of oil. This oil can be chosen from the group containing the traditional edible oil such as soybean oil, corn oil, sunflower oil or any other the or triglycerides, contains fatty acids short chain, such as miglyol, or white oil, such as paraffin oil or mineral oil consisting of a long hydrocarbon chain. Preferably the specified oil is sunflower oil, partially hydrogenated or rich in oleic acid.
Preferably the specified oil is injected in the following proportions relative to the substrate: undecylenoyl acid/oil = £ undecylenate acid - ¾ oil or 1/3 undecylenate acid - oil 2/3 or 1/2 undecylenate acid - 1/2 butter.
Mainly the specified oil is introduced into the fermentation medium with a flow rate of from 0.4 to 4.0 g/l/h, preferably from 0.5 to 3 g/l/h and very preferably from 0.75 to 2.5 g/l/h.
As any microorganisms in the conditions of forced bioconversion, Minety need a carbon source for their energy needs, as, for example, the source of glucide, preferably glucose or maltose. In the process of bioconversion is preferred intake of glucose. Therefore, in accordance with another preferred embodiment of the invention, glucose or maltose are entered simultaneously in undecylenoyl acid and oil. Primarily indicated the glucose and the specified maltose introduced into the fermentation medium with a flow rate of less than 1 g/l/h, preferably m is her 0.75 g/l/h and very preferably less than 0.5 g/l/h.
In accordance with another preferred embodiment of the invention the introduction of the specified undecylenate acid specified oil and possibly the indicated glucose or maltose in the medium fermentation is carried out continuously for 5-96 hours, preferably 24 to 72 hours and more preferably 24 to 48 hours.
In accordance with another preferred embodiment of the bioconversion according to the invention is carried out at a temperature from 25°C to 35°C, preferably from 27°C to 30°C.
In accordance with another preferred embodiment the pH of the fermentation medium is from 5 to 8, preferably 5.5 to 7.5, and more preferably from 6 to 7. In the framework of the invention regulate pH using sodium hydroxide (e.g., sterile NaOH5N or other Foundation, traditionally used for fermentation (Na4OH, KOH, etc)).
In the process of bioconversion is necessary intensive mixing, and good aeration of the fermentation medium. In the fermenter agitation is carried out using multiple blades. Mainly depending on the size of the fermenter, for example, if the digester has a volume of 6 liters, the need for appropriate mixing according to the invention is from 200 to 1200 rpm./min and preferably from 300 to 900 rpm./minutes with regard to aeration, it can become is determined by air flow at the level of the blades. Mainly proper aeration of the fermentation medium according to the invention is less than or equal to 1 vvm, preferably ranges from 0.2 to 0.8 vvm, and more preferably from 0.3 to 0.7 vvm.
In accordance with another preferred embodiment of the bioconversion of the invention is stopped by adding acid in the culture medium. The specified acid can be selected from the group consisting of phosphoric acid, sulfuric acid, hydrochloric acid or citric acid. Preferably, this acid is phosphoric acid or citric acid.
At the stage a) extraction of the 9-mission-2-it can be done by hydrodistillation or extraction with a solvent selected from the group consisting of cyclohexane, methylcyclohexane, or ethyl acetate. Preferably the solvent used cyclohexane or a mixture of cyclohexane and ethyl acetate.
Clean 9-mission-2-it can then be made by obessmolivanie with subsequent fractional distillation.
This method of extraction/purification allows, therefore, to obtain a molecule with a purity of more than 98% with the yield of extraction of about 75-85%.
The output of bioconversion 9-mission-2-in relation to the substrate, undecylenoyl acid is from about 25 to 35%.
The invention also relates to 9-mission-2-he obtained in this way, as described to enter the.
The object of the invention is the use of any 9-mission-2-he (CAS 35194-30-0 in perfumery and as a food flavoring, in particular, to obtain spirits, aromatic, cosmetic or food compositions or food supplements.
In a preferred embodiment, the object of the invention is the use of any 9-mission-2-he received a manner such as described above, in perfumery and as a food flavoring, in particular, to obtain spirits, aromatic, cosmetic or food compositions or food supplements.
In the context of the present invention, the term perfumery means not only perfumery in the usual sense, but also other areas in which the smell of the product is important. Speech, for example, you may go, but this is not a limitation, of perfumery compositions in the usual sense of the term, such as perfume bases and concentrates, colognes, toilet water, perfumes and similar products; topical composition - in particular, cosmetics, such as creams for face and body, talc, oils, hair shampoos, hair lotions, ointments, salts and bath oils, shower gels and bath, toilet soap, funds from perspiration and deodorants for body lotions and shaving creams, Soaps, creams, toothpaste, mouthwash for mouth, creams and similar products; the sanitary-hygiene, such as softeners, detergents, cleansers, deodorants for premises, aerosols and similar products.
The term odorant is used to refer to compounds which distributes the smell.
Under food flavouring understand any use of the compounds according to the invention to flavour any food, liquid or solid, for the food of man or animals, in particular, beverages, dairy products, ice cream, candy, chewing gum, and so on, as well as for flavoring tobacco products.
9-mission-2-it is also applicable in perfume compositions or food aromatic compositions for imparting notes of exotic, flower or fruit, particularly pineapple, pear, passion fruit exotic fruit. Depending on the application used, the amount of this molecule 9-mission-2-it is determined by a specialist in this field.
Preferably 9-mission-2-it will be used in amounts from 0,00025% to 30%, preferably from 0,0025% to 20%, more preferably from 0.025 to 10% of the mass. in relation to the total weight of the composition in which it is present. He cannot enter into the composition of solid or liquid compositions and, in particular, in the composition of gels, creams, ointments and/or sprays.
9-mission-2-it can also be used in the composition, which is itself the soul is the or in compositions in which the flavouring is used to hide or neutralize some of the odors.
Other aspects, objects, advantages and features of the invention are presented in the following description, having no limiting character, in which the described variants of the preferred embodiment and the invention by the following examples.
In Fig.1 shows the bioconversion of undecylenoyl acid to 9-mission-2-it.
Seeded strains of Mortierella isabellina (A), Aspergillus oryzae (b) and Penicillium roquefortii (C) [source = tube, frozen at -80°C] on agribiology malt and incubated at 27°C (case A) or 30°C (if b and C) within 72 hours.
Sow pre-culture in 3.5 liters of the medium malt in the fermenter with a volume of 6 l:
Extract of malt: 50 g/l
Yeast extract: 7,5 g/l
Incubated for 24 hours indicated strain at 27°C, with stirring at 500 rpm./min and aeration, corresponding to 0.3 vvm.
Aspergillus oryzae (B)
Incubated for 24 hours indicated strain at 30°C, under stirring speed of 600 rpm./min and aeration, corresponding to 0.5 vvm.
Penicillium roquefortii (C)
Incubated for 24 hours indicated strain at 30°C, under stirring speed of 600 rpm and aeration, corresponding to 0.3 vvm.>
Dry weight of mycelium after culturing each strain is approximately 9 to 11 g/l
Then undecylenoyl acid serves at a rate of 0.15 g/l/h for 6 hours, then with a flow rate of 0.33 g/l/h for 72 hours: i.e., in the amount of 25 g/L. This undecylenoyl acid serves in a mixture with hydrogenated sunflower oil (1/3 acid-2/3 oil; this oil is, thus, served with a flow rate of 0.6 g/l/h, then 1 g/l/h. In parallel continuously served the glucose consumption of 0.36 g/l/hour for 72 hours. pH is adjusted corresponding to 6.5 during the entire duration of the conversion with 5N NaOH. Increase the speed to 900 rpm./min and Mortierella isabellina aeronaut with a flow rate of 1 vvm, and Aspergillus oryzae or Penicillium roquefortii of 0.3 vvm. Conversion continued for 72 hours.
In non-optimized conditions, the yield of 9-mission-2-he is from 5 to 7 g/L.
Example 2: Conversion from Aspergillus oryzae ()
Sow Aspergillus oryzae [source = tube, frozen at -80°C] on agribiology malt and incubated at 30°C for 72 hours.
Sow pre-culture in 3.5 liters of the medium malt (the same concentration as in example 1).
Incubated at 30°C, 600 rpm./min and 0.5 vvm air free pH within 24 hours. Get the weight in dry matter 11 g/L.
Then undecylenoyl acid served with a flow rate of 0.5 g/l/h for 6 hours, then with a flow rate of 0.9 g/l/h for 48 hours: so what. in the amount of 46 g/L. This undecylenoyl acid serves in a mixture with hydrogenated sunflower oil (1/4 acid-3/4 oil; this oil is, thus, served with a flow rate of 1.5 g/l/h, then 2.7 g/l/h. In parallel continuously served the glucose consumption of 0.36 g/l/h for 48 hours. pH is adjusted corresponding 6 throughout the duration of the conversion with NaOH5N. Increase the speed to 900 rpm./min and aeronaut with a flow rate of 0.3 vvm. Conversion continue within 48 hours.
Exit 9-mission-2-it is 16.5 g/l, i.e. the output of the bioconversion is 35%.
Example 3: ConversionMortierellaisabellina (A)
Sow Mortierella isabellina [source = tube, frozen at -80°C] on agribiology malt and incubated at 27°C for 72 hours.
Sow the previous pre-culture in 3.5 liters of the medium malt (the same concentration as in example 1).
Incubated for 24 hours at 27°C, stirring, amounting to about 600./min, and aeration, corresponding to 0.5 vvm. Get the weight in the dry matter of 11.3 g/l
Then undecylenoyl acid served with a flow rate of 0.27 g/l/h for 6 hours, then consumption 0,477 g/l/h for 48 hours: i.e. a total of 46 g/L. This undecylenoyl acid serves in a mixture with hydrogenated sunflower oil (1/4 acid-3/4 oil; this oil is, thus, served with a flow rate of 0.8 g/l/h, then 1,43 g/l/h. Parallel continuous the EIT serves glucose with a flow rate of 0.32 g/l/h for 48 hours. pH is adjusted corresponding to 7.5 during the entire duration of the conversion with NaOH5N. Increase the speed to 900 rpm./min and aeronaut with a flow rate of 1 vvm. Conversion continue within 48 hours.
Exit 9-mission-2-he is 8 g/l, i.e. the output of the bioconversion is 33%.
Example 4: ConversionPenicillium roquefortii ()
Sow Penicillium roquefortii [source = tube, frozen at -80°C] on agribiology malt and incubated at 30°C for 72 hours.
Sow the previous pre-culture in 3.5 liters of the medium malt (the same concentration as in example 1).
Incubated for 22 hours at 30°C, stirring, amounting to about 600./min, and aeration, corresponding to 0.3 vvm. Get the weight in the dry matter of 10 g/L.
Then undecylenoyl acid re-served with a flow rate of 0.25 g/l/h for 6 hours, then with a flow rate of 0.45 g/l/h for 48 hours, then with a flow rate of 0.25 g/l/h for 24 hours: i.e., in the amount of 30 g/L. This undecylenoyl acid serves in a mixture with hydrogenated sunflower oil (1/3 acid-2/3 oil; this oil is, thus, served with a flow rate of 0.5 g/l/h, then 0.9 g/l/h, then, 0.5 g/l/h. In parallel continuously served the glucose consumption of 0.36 g/l/hour for 72 hours. pH is adjusted corresponding to 6.5 during the entire duration of the conversion with 5N NaOH. Increase the speed to 900 rpm./min and aeronaut with consumption ,6 vvm. Conversion continue within 48 hours.
Exit 9-mission-2-it is 10.5 g/l, i.e. the output of the bioconversion is 35%.
Example 5: extraction-cleanup
Is acidified to a pH of from 1.5 to 2 using 85% phosphoric acid. Enter solvent extraction with cyclohexane and stirred for 1 hour at room temperature. Centrifuged and Recuperat the organic phase. Determine the amount of ketone. The solvent is concentrated and thus receive the crude oily substance. Distilled in vacuum. Get "not" lactone extracted oil. Then cleanse, fractionary molecule in vacuum. Receive the product, purified more than 99%.
Example 6: Evaluation of 9-mission-2-he's in perfume
9-mission-2-he cleared 99%, tested on blotter and in solution (5% in ethanol): the head note in a strong, homogeneous, with notes of rose, aldehyde, acetate lemon grass, coriander leaves and sea notes. Heart has a pear notes.
Example 6: Evaluation as a food flavoring
9-mission-2-he cleared 99%, was tested in 8 memorial plaques in the mineral water: it has fruity notes of pear, Apple and pineapple and green and oily notes.
It is of interest to the complexity of natural recipes of exotic fruits, especially since there are not many natural molecules to achieve this is th goal. In particular, the tone of "Pineapple" 9-mission-2-he is a very attractive alternative to replace allelopathy, which is a molecule, most commonly used at present to get the tone type of the pineapple.
Example 8: Use in recipes as a food flavoring and in perfumery
Add 9-mission-2-it is in the aroma of exotic fruits allows you to get the typical "pineapple" tone. The following recipe equal weight replacement allelopathy commonly used in the formulation, 9-mission-2-he makes it very successfully to create the tone of "pineapple", which gives allylcapronate.
|Natural ethylcaproic||50 g|
|Natural maltol||20 g|
|Natural isoamylase||35 g|
|Natural utilizabilitate||37 g|
|Natural menthol codex||8 g|
|Natural acylmethyl-2-butyrate||75 g|
|Natural salivation||12 g|
|Natural ethyl butyrate||35 g|
|The purified camphor||2 g|
|Natural 9-mission-2-he||200 g|
|Natural ethyl alcohol||the number to 1000 g|
The use of 50 m D. this mixture in a weak juice drink gives it a very distinctive notes of "pineapple"; the use of 200 m D. this mixture makes sherbet typical "pineapple".
Also in the composition of natural food flavor 9-mission-2-it allows you to get interesting "pear" notes, for example, in the following formula:
|Natural hexanoic acid||from 1 to 10 g|
|Natural isoamyl alcohol||from 20 to 60 g|
|Natural hexyl alcohol||10 from 50 g|
|Natural ethyl kapat||from 20 to 80 g|
|from 50 to 150 g|
|Natural isoamylase||from 50 to 150 g|
|Natural propyl||from 50 to 150 g|
|Natural exilerated||from 50 to 150 g|
|Natural butyl acetate||from 50 to 150 g|
|Natural ethyl acetate||from 50 to 150 g|
|Natural 9-mission-2-he||from 20 to 50 g|
|Propylenglycol||the number to 1000 g|
The use of 100 meters D. this mixture in poorly podkamenno (5%) and weakly acidified beverage of 0.1% citric acid) gives it a very characteristic "pear" notes, even if 9-mission-2-it is found in very small quantities (2 to 5 M. D.).
In the perfume part 9-mission-2-it allows you to reinforce note floral, green, violet," and "aldehydic fruity shampoo, for example, in the following formula, which uses 1% 9-mission-2-he that makes a fragrance note of shampoo more complex and more intense:
|Phenethyl alcohol||100 g|
|Gamma undecalactone||4 g|
|Ambrettolide CIS-isomer||10 g|
|Anisic aldehyde||25 g|
|Ethyl vanillin||2 g|
|Digidrummer jasmonate||120 g|
|2-Actilingua acid, methyl ester||12 g|
In the case of other applications in perfumery, namely as a cushioning means, the presence of 0.5% 9-mission-2-it gives feature and enhances the fruity note, "Apple" or "pear", odor softening means, as indicated in the following formula:
|Phenethyl alcohol||80 g|
|Exilarchy aldehyde||40 g|
|Aldehyde C12||2 g|
|Ambrettolide CIS-isomer||60 g|
|3A,4,5,6,7,7 and hexahydro-4,7-methane-1H-indenify ether acetic acid||65 g|
|Essential oil patchouli||30 g|
|Cyclohexyl salicylate||100 g|
|Ambroxan 5% DPG||6 g|
|Essential oil galbani 10% DPG||10 g|
|Indole 10% DPG||8 g|
|9-mission-1-ol 10% DPG||10 g|
Finally, in the case of cream, 9-mission-2-it gives the last pronounced floral notes, for example, the following use:
|Phenethyl alcohol||65 g|
|Gamma undecalactone||6 g|
|Aljamal the glycolate||20 g|
|Anisic aldehyde||10 g|
|Hydrometer jasmonate||130 g|
|CIS-3 hexenol||2 g|
|Alpha ionon||23 g|
|Beta ionon||4 g|
|Essential oil patchouli||11 g|
|Gamma nonalactone||27 g|
|Hexenal CIS-3-salicylate||35 g|
|Van is Lin||2 g|
|Dimethylphenylcarbinol butyrate||1 g|
In the presence of 9-mission-2-cream is mainly the parent note with a more intense green and fresh notes.
Example 9: the toxicity Tests
The molecule was tested on 3 rabbits in accordance with the experimental Protocol established by the OECD guideline 404 N dated April 24th2002” and test “method B. 4 of the Directive N 2004/73/EC): Acute dermal irritation.
From this it follows that the 9-mission-2-it is not irritating and it does not require any symbol or phrase, warning about the risk (in accordance with EEC Directives 67/548/59 and 99/45).
The molecule was tested on 3 rabbits in accordance with the experimental Protocol established by the OECD guideline 405 N dated April 24th2002” and test “method B. 5 of the Directive N 2004/73/EC): Acute eye irritation.
It also follows that 9-mission-2-it is not irritating and it used what I don't need any symbol or phrase, a warning about the risk (in accordance with EEC Directives 67/548/59 and 99/45).
Finally, the value obtained during the test LD50 (acute oral toxicity (cf. the same Directive), namely 2500 mg/kg, allows to classify molecules as non-hazardous.
In connection with the security of the molecule, thus, it was found that this molecule can be used in recipes and in connection with the fact that her presence is established in mango and pineapple, can be used in the food and perfume industry as flavouring under the banner of "natural".
Although the present invention has been described above by way of examples of preferred variants of its implementation, it can be changed without deviating from the spirit and nature of the invention, such as defined in the attached claims.
1. A method of obtaining a 9-mission-2-it is characterized by bioconversion undecylenate acid using a mold, which belongs to the family Aspergillaceae or Mortierellaceae, and this method includes the following stages:
a) culturing a specified mold;
b) adding undecylenate acid as a substrate in a fermentation medium with a flow rate of from 0.1 to 0.9 g/l/h in the presence of oil, where the specified oil selected from the group consisting of traditional edible oils or triglycerides containing fatty acids short chain or alternative, white oil;
(C) the bioconversion of the substrate in the 9-mission-2-it,
d) extraction and purification 9-mission-2-it.
2. The method according to p. 1, characterized in that the mould belongs to the genus Aspergillus or the genus Penicillium.
3. The method according to p. 1, characterized in that the mould belongs to the genus Mortierella.
4. The method according to p. 1, characterized in that the stage of bioconversion includes the first phase of adaptation of the mould, during which the specified undecylenoyl acid is injected with a flow rate of from 0.1 to 0.5 g/l/h, then the second phase, during which the specified undecylenoyl acid is injected with a flow rate of from 0.25 to 0.9 g/l/h.
5. The method according to p. 4, characterized in that the duration of the specified phase adaptation is less than 20 hours, preferably less than 12 hours and very preferably less than 6 hours.
6. The method according to p. 1, characterized in that the oil is introduced into the fermentation medium with a flow rate of from 0.4 to 4.0 g/l/h.
7. The method according to p. 1, wherein the glucose or maltose injected simultaneously with undecylenate acid and oil.
8. The method according to p. 7, characterized in that the specified glucose or specified maltose is introduced into the fermentation medium with a flow rate of less than 1 g/l/h, preferably less than 0.75 g/l/h and very preferably less than 0.5 g/l/h.
9. The method according to p. 1, characterized in that the introduction of the specified undecylenate acid specified oil and may specify the Oh of glucose or maltose in the medium fermentation carried out continuously for 5-96 hours.
10. The method according to p. 1, wherein the bioconversion is carried out at a temperature from 25°C to 35°C, preferably from 27°C to 30°C.
11. The method according to p. 1, characterized in that the pH of the fermentation medium is from 5 to 8.
12. The method according to p. 1, characterized in that the aeration of the fermentation medium is less than or equal to 1 vvm.
13. The method according to p. 1, wherein the bioconversion is stopped by adding to the culture medium acid selected from the group consisting of phosphoric acid, sulfuric acid, hydrochloric acid or citric acid.
SUBSTANCE: method comprises growing cells of the strain of bacteria Rhodococcus rhodochrous RNCIM AC-1898. The cells are precipitated and washed three times with phosphate-alkaline buffered solution. It is re-suspended in the same solution and the optical density of the cell suspension is adjusted to a predetermined value, and betulin is added in the dimethyl sulphoxide solution at a concentration of from 0.5 to 3.0 g/l followed by biotransformation of betulin to betulon. And the process of biotransformation takes 24-48 hours at the optical density of the cell suspension of 2.0-2.6 (dry weight of biomass of 7-10 g/l).
EFFECT: invention enables to increase the yield of betulon.
2 cl, 2 dwg, 4 ex
SUBSTANCE: method is proposed to produce difunctional alkanes in a recombinant host cell from initial material of carbohydrates in host cells.
EFFECT: invention may be used to produce compounds using microorganisms for production of microorganisms for production of chemical substances, production of which by conventional methods is complicated and expensive.
7 cl, 5 dwg, 5 tbl
SUBSTANCE: invention is related to biotechnology and may be used to produce butanol, acetone and ethanol. Strain of bacteria Clostridium acetobutylicum is deposited in All-Russia collection of microorganisms, IBPM named after G.K. Skryabin under number B-2531D and is a producer of butanol, acetone and ethanol.
EFFECT: invention makes it possible to increase yield of butanol, acetone and ethanol and to expand base of raw materials for their production.
1 tbl, 3 ex
SUBSTANCE: strain Trametes versicolor is proposed, used for production of antifungal agents against fungi of the genus Penicillium. The strain is deposited in the RCIM under the number F-1024.
EFFECT: strain has high chitinase and fungicidal activity.
4 dwg, 1 tbl, 5 ex
SUBSTANCE: invention relates to biotechnology and can be used in agriculture. The Trichoderma harzianum Rifai strain is deposited in the Russian National Collection of Industrial Microorganisms under registration number VKPM F-180. The strain is capable of producing L-lysine-alpha-oxidase and can be used particularly as an inhibitor of cucurbit bacterial spot caused by Acidovorax citrulli bacteria.
EFFECT: invention reduces loss of cucurbit crop.
SUBSTANCE: method for obtaining proteinase - protein C activator of blood plasma provides for solid-phase cultivation of fungus strain Aspergillus ochraceus BKM F-4104D on a substrate. The substrate contains the following components at a certain ratio: a solid substance or a carrier, glucose, amylum, hydrolysate of fish flour, peptone, NaCl, KH2PO4, MgSO4·7H2O, and water. As a solid substance or a carrier, vermiculite, polyurethane foam, silica gel or wheat middlings are used. Proteinase activity is up to 300 U/ml.
EFFECT: invention allows obtaining a target product with improved activity.
5 tbl, 15 ex
SUBSTANCE: group of inventions relates to biotechnology. A method of obtaining a granulated product includes growing filamentous fungi of a Monilialeae family, preferably Arthrobotrys conoides Dreschsler in a suitable liquid culture medium. The obtained culture of the filamentous fungus is mixed with at least one type of modified starch and starch flour, with the modified starch and modified flour being present in weight ratios, constituting from 30:70 to 60:40. Filling agents and in case of necessity nutritional substances are added to the obtained mixture with obtaining a paste. The granulation of the paste is carried out. The obtained granules are dried until the moisture level lower than 13% is achieved, preferably to the level constituting from 9 to 10%. The granulated product, obtained by the method described above, is used for the application in a pesticidal composition.
EFFECT: group of inventions provides obtaining the target product with high dispersability and a high percent of survival of the filamentous fungi propagules.
9 cl, 1 dwg, 4 ex
SUBSTANCE: group of inventions relates to biotechnology. The method of obtaining a biological preparation for the protection of plants against phytopathogens and nematodes based on a genus Trichoderma fungus strain is realised by preparation of an inoculation material of the fungus strain, making the preparation in a liquid or friable form based on the inoculation material, and mixing the preparation with a mineral, organic or bacterial fertiliser.
EFFECT: biological preparation, resulting from the method, possesses the high antagonistic activity with respect to a wide spectrum of phytopathogens and nematodes.
6 cl, 5 ex
SUBSTANCE: group of inventions relates to biochemistry. Claimed is a device for surface growing of a microorganism on a liquid culture medium. The device includes a dish with a drain pan for growing of the microorganism, provided with a transport net freely dropped on its bottom. The transport net is made from a neutral material and is fastened by its ends to drive drums, which ensure lifting of the transport net from the dish bottom. The device also contains a reception device with a valve for the connection to a mixer-doser and supply of the liquid culture medium mixture with mother culture into the dish, rotary flaps, placed on butt ends of the dish for the regulation of air exchange in the dish, a drain device for pouring out the culture liquid, a pipe branch for the supply of a drying agent under the net, a sampling instrument and removable colour filters on lamps for the regulation of illumination, placed on side walls of the dish, a lid, which hermetically closes the dish, made with a possibility of its opening and/or removal, equipped with injectors for spraying the SAS solution above the surface of biomass growing in the dish and washing heads for the realisation of sanitary processing of the device. Also claimed is a method of surface growing of a microorganism on a liquid culture medium with the application of the claimed device.
EFFECT: claimed inventions ensure growing of microorganisms on the liquid culture medium with the minimal application of manual labour.
7 cl, 1 dwg, 2 ex
SUBSTANCE: wound-healing agent is a concentrate of the culture liquid of strain Trichoderma harzianum Rifai deposited in the Russian National Collection of Industrial Microorganisms under the number of RNCIM: F-180, as a producer of L-lysine of alpha-oxidase, and can be applied as a wound-healing agent for skin lesions.
EFFECT: invention enables to expand the range of means that provide wound healing of skin lesions.
1 tbl, 2 ex
SUBSTANCE: invention relates to the use of the concentrate of the culture liquid of the strain Trichoderma harzianum Rifai, deposited in the Russian National Collection of Industrial Microorganisms under the number of RNCIM F-180 as an inhibitor of Andis virus of potato mottling.
EFFECT: invention enables to reduce losses of potato from the plant infection with the Andis mottling virus.
SUBSTANCE: nutrient medium for growing filamentous fungi-dermatophytes from clinical material comprises glucose, bacteriological agar, meat peptone, casein hydrolyzate, yeast extract, sodium chloride, sodium carbonate, L-cysteine, thioglycolic acid and distilled water in a predetermined ratio of components.
EFFECT: invention enables to reduce time of growing filamentous fungi-dermatophytes from clinical material.
1 tbl, 2 dwg, 4 ex
SUBSTANCE: invention relates to biotechnology. The method involves treating the sensitive layer of a biosensor with solution of a common fraction of protease of hepatopancreas of king crab in a buffer comprising tris-HCl 50 mM, CaCl2 3 mM, NaCl 100 mM, pH 8.0 at solution temperature of 35-40°C. The process is carried out in multiple successive steps with a solution of a solution of a common fraction of protease of hepatopancreas of king crab in said solvent at temperature of 35-40°C followed by exposure. The sensitive layer of the biosensor is successively washed with dodecyl sulphate dissolved in the same buffer in concentration of 0.1% and then with water at each step. The treatment process is carried out three times.
EFFECT: invention reduces the duration of the process.
FIELD: biotechnology, in particular reagent for structural protein hydrolysis.
SUBSTANCE: method for production of protheolytic reagent includes cultivation of producer strain selected from dermatophyte of species Trichophyton mentagrophytes, Trichophyton verrucosum, Trichophyton equinum, Microsporum canis, Microsporum gypseum, preferably from fungus strains capable to produce complex of structural protein proteinases (scleroproteases) with total protheolytic activity at least 0.7 U/mg including collagenase 0.1-4.5 U/mg; keratinase 0.1-0.5- U/mg; elastase 0.5-1.9 U/mg. Cultivation is carried out, for example, in wort agar-agar or in wort broth for 20-24 days.
EFFECT: scleroproteases with improved specific activity; method for protheolytic cleavage of specific substrates (scleroproteins) with increased rate and depth.
2 dwg, 12 ex
FIELD: microbiological industry.
SUBSTANCE: invention relates to nutrient media used for culturing producers of carotene. Invention proposes nutrient medium containing barley flour, soybean meal, potassium dihydrogen phosphate, sunflower oil, vitamin B1, β-ionone and tap water being components are taken in the following ratio, wt.-%: barley flour, 3.0-4.0; soybean meal, 4.0-4.7; sunflower oil, 3.8-4.0; potassium dihydrogen phosphate, 0.04-0.05; vitamin B1, 0.0002-0.0005; β-ionone, 0.098-0.099, and tap water, the balance. The proposed nutrient medium is low-priced and enhances the biosynthetic ability of producer of carotene.
EFFECT: valuable properties of nutrient medium.
3 tbl, 3 ex
FIELD: biotechnology, microbiology.
SUBSTANCE: invention relates to producing ethyl alcohol or fodder for monogastral animals. The polyenzyme product with glucoamylase, proteolytic and xylanase activities is prepared by fermentation of wheat bran using microorganism Aspergillus niger. Indicated glucoamylase, proteolytic and xylanase activities have the following minimal values: glucoamylase activity - at least 100 U/g of dry matter; proteolytic activity - at least 100 U/g of dry matter; xylanase activity - at least 100 U/g of dry matter under condition that glucoamylase activity has to be at least 750 U/g of dry matter and/or xylanase activity has to be at least 300 U/g of dry matter. Invention provides enhancing the soluble nitrogen content in wort after saccharification, reducing viscosity and effectiveness in using.
EFFECT: improved preparing method, valuable properties of hydrolyzed bran.
14 cl, 10 tbl, 7 ex
FIELD: biotechnology, microbiology.
SUBSTANCE: invention proposes a method for preparing exopolysaccharide by culturing microorganisms in nutrient medium containing one or more carbon source assimilated by microorganisms and caruba seeds fraction as nitrogen organic source. Applying the proposed method provides preparing exopolysaccharide eliciting improved organoleptic, sensor and visual properties. Invention can be used in building, paper, textile, cosmetic, food, oil output industry and agriculture.
EFFECT: improved preparing method.
15 cl, 4 ex
FIELD: agriculture, fruit storage.
SUBSTANCE: the present innovation deals with technology to treat fruit before their laying for storage. For the purpose to decrease the loss from microbial spoiling and increase the terms of storage fruit essential oils and epiphytic substances of cuticular layer of plant raw material should be applied onto the surface from mycella obtained after extracting Saprolegnia parasitica micromycete biomass with nonpolar extragent in supracritical state.
EFFECT: higher efficiency of protection.
FIELD: food industry, confectionary industry.
SUBSTANCE: the suggested concentrate should be prepared due to pressing the juice under aseptic conditions out of sugar beet prepared, deodorated and sterilized with carbon dioxide in supracritical state in the field of ultrasound fluctuations, cultivating upon residues of mycellial fungi of Trichoderma and Aspergillus species of citric acid fermentation, separating culture liquid, its mixing with juice and introducing liquid ammonia into the blend along with supracritical CO2-extract out of Mortierella nigrescens micromycete biomass further extracted according to the preset technique to obtain a solid residue treated with liquid ammonia followed by concentrating the blend, treatment of concentrate with liquid carbon dioxide, mixing Mortierella nigrescens micromycete biomass with a treated solid residue and heating the mixture up to 60 C, not less. The innovation enables to obtain a concentrate of improved structure-forming properties and increased thermal stability.
EFFECT: higher efficiency of manufacturing.