CN101731568A - Method for preparing high-salt dilute soy by adopting immobilized cell fermentation - Google Patents
Method for preparing high-salt dilute soy by adopting immobilized cell fermentation Download PDFInfo
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Abstract
The invention discloses a method for preparing a high-salt dilute soy by adopting immobilized cell fermentation, which comprises the following steps of: 1. preparing a soy by using a traditional high-salt dilute fermentation method and preparing a soy stock solution through fermentation for 60-90 days for standby; 2. fetching four strains of activated torulopsis, zygosaccharomyces rouxii, shengxiangsaite candida and soy tetrads to respectively prepare immobilized cell particles; 3. compounding the prepared immobilized cell particles of the four strains of the torulopsis, the zygosaccharomyces rouxii, the shengxiangsaite candida and the soy tetrads in the step 2 to obtain mixed immobilized cell particles; then inoculating the mixed immobilized cell particles into the high-salt dilute fermentation soy stock solution obtained in the step 1 for fermentation; and 4. after fermentation is finished, separating the maturely fermented high-salt dilute fermentation soy and the mixed immobilized cell particles to obtain a finished product of the high-salt dilute fermentation soy. The method not only can enhance the taste of the high-salt dilute soy, but also shortens the period of soy fermentation and overcomes the defects of long fermentation period, low equipment utilization ratio and higher production cost of the high-salt dilute fermentation method.
Description
Technical field
The present invention relates to a kind of method for preparing high-salt dilute soy, relate in particular to a kind of method that adopts immobilized cell fermentation to prepare high-salt dilute soy.
Background technology
Soy sauce is a kind of time-honored flavouring, and the brewing method of at present domestic soy sauce mainly contains two kinds of high-salt dilute fermentation method and low-salt solid-state fermentation methods.Low-salt solid-state fermentation method fermentation period is short, but product special flavour can not show a candle to the high-salt dilute fermentation method.The high-salt dilute zymotechnics is to be major ingredient with soya bean and flour, and through microbial fermentation kojis such as aspergillus oryzaes, the fermentation process that utilizes the natural solarization of high-salt dilute to reveal is brewageed and formed.The biochemistry that the sweat of high-salt dilute zymotechnique mainly contains two stages changes: the phase I is at the fermentation initial stage, promptly preceding 60～90 days of fermentation, mainly be the protein in the raw material to be resolved into amino acid (main component in the soy sauce) and reduced sugar by protease, carbohydrase that aspergillus oryzae is produced in the Daqu preparation process, form the delicate flavour material of soy sauce, also can produce the composition that has sauce fragrant breeze flavor on a small quantity simultaneously; Second stage is in the fermentation later stage, mainly constantly produce organic compounds such as ethanol, lactic acid during the fermentation by microorganisms such as some saccharomycete, lactic acid bacterias, biochemistry by complexity changes main flavor in the generation soy sauce, form the exclusive mellow sauce fragrant breeze flavor of high-salt dilute soy, as ethanol, lactic acid, 4-vinyl guajacolum etc.Therefore, high-salt dilute fermentation needs 4-6 month time could form the mellow sauce fragrant breeze of soy sauce to distinguish the flavor of, but it is long to have a fermentation time, the shortcoming that utilization rate of equipment and installations is low.
Conventional method is primarily aimed at the research that promotes flavor of soy sauce and controls key process parameter etc. in raw material proportioning, raw material specially treated, bacterial classification transformation, additive and sweat.The development in several thousand of sauce fermentation technology, though make the research of conventional method certain help be arranged to the local flavor that promotes soy sauce, but continuous upgrading along with technology, technological parameter traditional in the sauce fermentation is definite substantially, amplitude of variation is less, and is very limited to the space that promotes flavor of soy sauce; Though the proper extension fermentation period can improve its product special flavour, fermentation period surpassed after 120-150 days, the DeGrain that local flavor improves, and prolong the turnover utilization that fermentation period has limited equipment, improved production cost virtually; Secondly, in the long process of sauce fermentation, its product special flavour is subjected to difference, the various places natural conditions of raw material, many-sided factor affecting such as variation of climatic factor, the difficult control of the stability of product special flavour.
Immobilized cell fermentation is the biotechnology of develop rapidly in recent years, it is microbial cell to be adhered to be fixed on certain carrier, the unit volume bacteria containing amount substantially exceeds conventional fermentation bacteria containing amount in the reactor, thereby can improve fermentation rate and shorten fermentation period.
Japan begins one's study in nineteen eighty-two immobilized cell fermentation is used for soy sauce brewing, and the researcher has done the test of a large amount of immobilization salt tolerant yeast bacterium and lactic acid bacteria thereafter.But all being that to adopt the less salt zymotic fluid be substrate, is carrier with sodium alginate, polyvinyl alcohol or ceramic, respectively fixedly salt tolerant yeast bacterium and lactic acid bacteria.Show according to foreign study: by immobilized cell technology, immobilized cell concentration and metabolite all improve than free cell, the physico-chemical analysis value of immobilized cell fermentation product meets the traditional fermentation standard for soy sauce, but sensory evaluation does not still reach the level of traditional high-salt liquid-state brewing soy sauce.At present also there is not manufacturing enterprise can realize the commercial application of immobilized cell technique in the actual production process of high-salt liquid-state brewing soy sauce.
Summary of the invention
The objective of the invention is to overcome the deficiencies in the prior art, a kind of method that adopts immobilized cell fermentation to prepare high-salt dilute soy is provided, this method can not only promote the high-salt dilute soy local flavor and shorten the cycle of sauce fermentation, and can also overcome the shortcoming that high-salt dilute fermentation method fermentation period is long, utilization rate of equipment and installations is low, production cost is higher.
The objective of the invention is to realize by following technical measures: a kind of method that adopts immobilized cell fermentation to prepare high-salt dilute soy, it may further comprise the steps:
(1) prepares soy sauce with traditional high-salt dilute fermentation method, make through the soy sauce stoste of fermentation in 60-90 days standby;
(2) learn from else's experience activation torulopsis, Lu Shi yeast, give birth to fragrant Saite Candida and four kinds of bacterial classifications of soy sauce tetrads are made the immobilized cell particle respectively;
The immobilized cell particle of (3) get the torulopsis that makes in the step (2), Lu Shi yeast, giving birth to fragrant Saite Candida and four kinds of bacterial classifications of soy sauce tetrads is according to mass ratio 0～3: carry out composite obtain mixed immobilization cell particle at 0～3: 3～5: 0.1～1; Be that 10-30% (W/V) is seeded to mixed immobilization cell particle in the high-salt dilute fermented sauce stoste of step (1) gained then with the inoculum concentration, continuously fermented 15-24 hour at temperature 28-33 ℃;
(4) after the fermentation ends, with after the high-salt dilute fermented sauce of this fermenting-ripening and the immobilized cell separate particles high-salt dilute fermented sauce finished product.
Torulopsis, Lu Shi yeast, living fragrant Saite Candida and the activation of four kinds of bacterial classifications of soy sauce tetrads and the preparation of their immobilized cell particle adopt existing method all can realize purpose of the present invention in the step of the present invention (2).And as a kind of embodiment wherein, the immobilized cell particle can make by following steps:
The learn from else's experience torulopsis, Lu Shi yeast of activation, the bacterium liquid of giving birth to fragrant Saite Candida and four kinds of bacterial classifications of soy sauce tetrads is mixed with into cell suspending liquid with the sodium alginate glue by 1: 15～25 volume ratio respectively, then cell suspending liquid being instilled into mass percent concentration is in 2% (W/W) calcium chloride solution, promptly obtains the immobilized cell particle of torulopsis, Lu Shi yeast, living fragrant Saite Candida and four kinds of bacterial classifications of soy sauce tetrads respectively.
The mass percent concentration of described sodium alginate glue is 1.5～3.5%; Described mass percent concentration is that the addition of calcium chloride solution of 2% (W/W) and the volume ratio of sodium alginate bacterium liquid are 1.5～3: 1 (V/V).
It is 10 that the immobilized cell particle of described torulopsis, Lu Shi yeast, living fragrant Saite Candida and four kinds of bacterial classifications of soy sauce tetrads all can require the average bacteria containing amount in the immobilized cell particle
6～10
8Individual/particle, can guarantee that each bacterial classification has suitable concentration in the sauce fermentation raw material, and guarantee that the sauce fermentation raw material can fully ferment, improved fermenting speed, obtain good ferment effect.
The present invention can add mass percent concentration in the high-salt dilute fermented sauce stoste of step (3) be the liquefaction syrup of 5～10% (W/W), its addition is 5～20% (W/W) of high-salt dilute fermented sauce stoste gross weight, for torulopsis, Lu Shi yeast, the fermentation of giving birth to fragrant Saite Candida and soy sauce tetrads provide enough nutrients, to promote the breeding growth of each bacterial classification, help producing effective flavor substance, make sense organ unique flavor, the sauce of the soy sauce that finally obtains aromatic thick.
The present invention can do following improvement: once stir every 30～60min in the process of continuously fermenting described in the step (3); Each time of stirring is 2～5min, and mixing speed can be in 80～150r/min scope.Stir during the fermentation, the bacterial classification in the fermentation materials evenly distributes with nutrient and can fully contact with fermenation raw liquid, and improves the efficient of dissolved oxygen, improves fermentation efficiency and effect.
In step of the present invention (4) the high-salt dilute fermented sauce of fermenting-ripening and immobilized cell separate particles adopt in the prior art effective separation method all can, comprise centrifugation, filtration etc.
The present invention compared with prior art has following beneficial effect:
(1) the present invention is on the production technology basis of traditional high-salt dilute soy, utilize immobilized cell technology, is that 60-90 days soy sauce stoste is fermented with the bacterial classification that mixes to fermentation time, improves the content and the ratio thereof of flavor substances such as alcohols in the soy sauce, phenols, ester class and heterocycle compound.After testing, utilize the soy sauce of this technology fermentation and soy sauce that 120-150 days traditional handicraft of need fermentation obtains to compare, the content of several main flavor such as its 1-octene-3-alcohol, ethanol, 4-vinyl guajacolum all is significantly improved, thereby promote high-salt dilute fermented sauce local flavor, obtain sense organ unique flavor, the aromatic thick soy sauce of sauce.
(2) production cycle of the present invention has shortened the production cycle of high-salt dilute fermented sauce greatly about 60～90 days, has improved the utilization rate of Zymolysis Equipment, has reduced production cost.
(3) after each bacterial classification among the present invention carries out cell fixation, can reclaim repeated use, comparatively stable to the control of local flavor during the fermentation.
The specific embodiment
Embodiment one:
Adopt immobilized cell fermentation to prepare the method for high-salt dilute soy:
(1) gets torulopsis, Lu Shi yeast, living fragrant Saite Candida respectively, be that the sucrose solution of 2% (W/W) activates under 30 ℃ of constant temperatures with mass percent concentration respectively, each barms vigor is fully recovered, spread cultivation then, centrifugal, stand-by with the sterilized water dilution, bacterial classification: sterilized water=1: 5 (W/V).
(2) soy sauce tetrads bacterial classification inoculation is activated more than 3 times in degreasing milk medium, spawn activity is fully recovered, it is centrifugal to spread cultivation then, stand-by with the sterilized water dilution, bacterial classification: sterilized water=1: 20 (W/V).
(3) the traditional high-salt dilute soy fermentation method of employing prepares the high-salt dilute soy stoste through fermenting 60 days, and is standby.
(4) preparation method of bacterial classification immobilization particle: with the torulopsis of step (1) and step (2) preparation, the Lu Shi yeast, to be 2% (W/W) sodium alginate glue with mass percent concentration respectively be mixed into cell suspending liquid by the volume ratio of 1: 20 (V/V) to the bacterium liquid of giving birth to fragrant Saite Candida and soy sauce tetrads, then cell suspending liquid being instilled into mass percent concentration is in 2% (W/W) calcium chloride solution, wherein the volume ratio of 2% calcium chloride solution and sodium alginate cell suspending liquid is 2: 1 (V/V), promptly obtains torulopsis respectively, the Lu Shi yeast, give birth to the pearl immobilization calcium alginate particle of fragrant Saite Candida and soy sauce tetrads.Prepare corresponding immobilized cell particle as stated above.The average bacteria containing amount that requires each immobilized cell particle is 10
6Individual/particle, then with torulopsis, Lu Shi yeast, give birth to fragrant Saite Candida and four kinds of immobilized cell particles of soy sauce tetrads by 1: 1: 3: the composite mixing of 0.5 mass ratio.
(5) get the high-salt dilute soy stoste of 100kg step (3) gained, add the 10kg mass percent concentration and be 10% liquefaction syrup, the mixed bacteria immobilized cell particle for preparing is inoculated into soy sauce stoste, inoculum concentration is 20% (W
Mixed bacteria immobilized cell particle/ V
High-salt dilute soy stoste), ferment then, under 28 ℃ of fermentation temperatures, continuously fermented 24 hours, stir 2min every 30min, mixing speed 120r/min.
(6) after the fermentation ends, the soy sauce of fermenting-ripening is separated with the immobilized cell particulate filter, obtain high-salt dilute soy.Immobilized cell particle after the recovery can ferment next time.
Composition comparative analysis: utilize the method for SPME-gas chromatography mass spectrometry that the soy sauce sample of traditional handicraft fermentation in 120 days is analyzed with the volatile flavor component of the soy sauce sample that utilizes the immobilized cell technology fermentation.Testing result shows: utilize the content raising 20% of the soy sauce sample of immobilized cell fermentation than the 1-octene-3-alcohol in the soy sauce sample of fermentation in 120 days, 4-vinyl guajacolum content has improved 30%, and total ester has improved 8%, and ethyl acetate has improved 5%.
Sense organ compares: 120 days traditional handicraft fermenting-ripening soy sauce samples are carried out subjective appreciation with the soy sauce sample that utilizes the immobilized cell technology fermentation.Subjective appreciation is the result show: the soy sauce sample that utilizes the flavor of soy sauce of the soy sauce sample of immobilized cell technology fermentation to ferment than 120 days traditional handicrafts has clear improvement, and sauce perfume (or spice) is more mellow.
Embodiment two:
Adopt immobilized cell fermentation to prepare the method for high-salt dilute soy:
(1) gets torulopsis, Lu Shi yeast, living fragrant Saite Candida respectively, be that the sucrose solution of 2% (W/W) activates under 30 ℃ of constant temperatures with mass percent concentration respectively, each barms vigor is fully recovered, spread cultivation then, centrifugal, stand-by with the sterilized water dilution, bacterial classification: sterilized water=1: 4 (W/V).
(2) soy sauce tetrads bacterial classification inoculation is activated more than 3 times in degreasing milk medium, spawn activity is fully recovered, it is centrifugal to spread cultivation then, stand-by with the sterilized water dilution, bacterial classification: sterilized water=1: 15 (W/V).
(3) the traditional high-salt dilute soy fermentation method of employing prepares the high-salt dilute soy stoste through fermenting 70 days, and is standby.
(4) preparation method of bacterial classification immobilization particle: with the torulopsis of step (1) and step (2) preparation, the Lu Shi yeast, giving birth to fragrant Saite Candida and soy sauce tetrads bacterium liquid is that the sodium alginate glue of 3% (W/W) is mixed into cell suspending liquid by the volume ratio of 1: 15 (V/V) with mass percent concentration respectively, then cell suspending liquid is instilled in the calcium chloride solution that mass percent concentration is 2% (W/W), wherein the volume ratio of 2% calcium chloride solution and sodium alginate cell suspending liquid is 1.5: 1 (V/V), promptly obtains torulopsis respectively, the Lu Shi yeast, give birth to the pearl immobilization calcium alginate particle of fragrant Saite Candida and soy sauce tetrads.The average bacteria containing amount that requires each immobilized cell particle is 107 a/particle, then pressing 1: 2: 4 torulopsis, Lu Shi yeast, living fragrant Saite Candida and four kinds of immobilized cell particles of soy sauce tetrads: the composite mixing of 0.3 mass ratio.
(5) get the high-salt dilute soy stoste of 100kg step (3) gained, adding the 13kg mass percent concentration is the liquefaction syrup of 8% (W/W), and the mixed bacteria immobilized cell particle for preparing is inoculated into soy sauce stoste, and inoculum concentration is 15% (W
Mixed bacteria immobilized cell particle/ V
High-salt dilute soy stoste), ferment then, under 30 ℃ of fermentation temperatures, continuously fermented 18 hours, stir 5min every 60min, mixing speed 100r/min.
(6) after the fermentation ends, the soy sauce of fermenting-ripening is separated with the immobilized cell particulate filter, obtain high-salt dilute soy.Immobilized cell particle after the recovery can ferment next time.
Composition comparative analysis: utilize the method for SPME-gas chromatography mass spectrometry that the soy sauce sample of traditional handicraft fermentation in 120 days is analyzed with the volatile flavor component of the soy sauce sample that utilizes the immobilized cell technology fermentation.Testing result shows: utilize the content raising 23% of the soy sauce sample of immobilized cell fermentation than the 1-octene-3-alcohol in the soy sauce sample of fermentation in 120 days, 4-vinyl guajacolum content has improved 28.74%, and total ester has improved 10%, and ethyl acetate has improved 5%.
Sense organ compares: 120 days traditional handicraft fermenting-ripening soy sauce samples are carried out subjective appreciation with the soy sauce sample that utilizes the immobilized cell technology fermentation.Subjective appreciation is the result show: the soy sauce sample that utilizes the flavor of soy sauce of the soy sauce sample of immobilized cell technology fermentation to ferment than 120 days traditional handicrafts has clear improvement, and sauce perfume (or spice) is more mellow.
Embodiment three:
Adopt immobilized cell fermentation to prepare the method for high-salt dilute soy:
(1) gets torulopsis, Lu Shi yeast, living fragrant Saite Candida respectively, be that the sucrose solution of 2% (W/W) activates under 30 ℃ of constant temperatures with mass percent concentration respectively, each barms vigor is fully recovered, spread cultivation then, centrifugal, stand-by with the sterilized water dilution, bacterial classification: sterilized water=1: 6 (W/V).
(2) soy sauce tetrads bacterial classification inoculation is activated more than 3 times in degreasing milk medium, spawn activity is fully recovered, it is centrifugal to spread cultivation then, stand-by with the sterilized water dilution, bacterial classification: sterilized water=1: 25 (W/V).
(3) the traditional high-salt dilute soy fermentation method of employing prepares the high-salt dilute soy stoste through fermenting 90 days, and is standby.
(4) preparation method of bacterial classification immobilization particle: with the torulopsis of step (1) and step (2) preparation, the Lu Shi yeast, giving birth to fragrant Saite Candida and soy sauce tetrads bacterium liquid is that the sodium alginate glue of 2% (W/W) is mixed into cell suspending liquid by the volume ratio of 1: 25 (V/V) with mass percent concentration respectively, then cell suspending liquid being instilled into mass percent concentration is in 2% calcium chloride solution, wherein the volume ratio of 2% calcium chloride solution and sodium alginate cell suspending liquid is 2: 1 (V/V), promptly obtains torulopsis respectively, the Lu Shi yeast, give birth to the pearl immobilization calcium alginate particle of fragrant Saite Candida and soy sauce tetrads.The average bacteria containing amount that requires each immobilized cell particle is 107 a/particle, then pressing 1: 3: 3 torulopsis, Lu Shi yeast, living fragrant Saite Candida and four kinds of immobilized cell particles of soy sauce tetrads: the composite mixing of 1 mass ratio.
(5) get the high-salt dilute soy stoste of 100kg step (3) gained, add the 20kg mass percent concentration and be 5% liquefaction syrup, the mixed bacteria immobilized cell particle for preparing is inoculated into soy sauce stoste, inoculum concentration is 25% (W
Mixed bacteria immobilized cell particle/ V
High-salt dilute soy stoste), ferment then, under 32 ℃ of fermentation temperatures, continuously fermented 15 hours, stir 3min every 30min, mixing speed 100r/min.
(6) after the fermentation ends, the soy sauce of fermenting-ripening is separated with the immobilized cell particulate filter, obtain high-salt dilute soy.Immobilized cell particle after the recovery can ferment next time.
Composition comparative analysis: utilize the method for SPME-gas chromatography mass spectrometry that the soy sauce sample of traditional handicraft fermentation in 120 days is analyzed with the volatile flavor component of the soy sauce sample that utilizes the immobilized cell technology fermentation.Testing result shows: utilize the content raising 20% of the soy sauce sample of immobilized cell fermentation than the 1-octene-3-alcohol in the soy sauce sample of fermentation in 120 days, 4-vinyl guajacolum content has improved 33.5%, and total ester has improved 8%, and ethyl acetate has improved 3%.
Sense organ compares: 120 days traditional handicraft fermenting-ripening soy sauce samples are carried out subjective appreciation with the soy sauce sample that utilizes the immobilized cell technology fermentation.Subjective appreciation is the result show: the soy sauce sample that utilizes the flavor of soy sauce of the soy sauce sample of immobilized cell technology fermentation to ferment than 120 days traditional handicrafts has clear improvement, and sauce perfume (or spice) is more mellow.
Embodiment four:
Adopt immobilized cell fermentation to prepare the method for high-salt dilute soy:
(1) gets Lu Shi yeast, living fragrant Saite Candida respectively, be that the sucrose solution of 2% (W/W) activates under 30 ℃ of constant temperatures with mass percent concentration respectively, each barms vigor is fully recovered, spread cultivation then, centrifugal, stand-by with the sterilized water dilution, bacterial classification: sterilized water=1: 5 (W/V).
(2) soy sauce tetrads bacterial classification inoculation is activated more than 3 times in degreasing milk medium, spawn activity is fully recovered, it is centrifugal to spread cultivation then, stand-by with the sterilized water dilution, bacterial classification: sterilized water=1: 20 (W/V).
(3) the traditional high-salt dilute soy fermentation method of employing prepares the high-salt dilute soy stoste through fermenting 80 days, and is standby;
(4) preparation method of bacterial classification immobilization particle: with the Lu Shi yeast of step (1) and step (2) preparation, to be 2% (W/W) sodium alginate glue with mass percent concentration respectively be mixed into cell suspending liquid by the volume ratio of 1: 20 (V/V) to the bacterium liquid of giving birth to fragrant Saite Candida and soy sauce tetrads, then cell suspending liquid being instilled into mass percent concentration is in 2% (W/W) calcium chloride solution, wherein the volume ratio of 2% calcium chloride solution and sodium alginate cell suspending liquid is 3: 1 (V/V), promptly obtains the Lu Shi yeast respectively, give birth to the pearl immobilization calcium alginate particle of fragrant Saite Candida and soy sauce tetrads.The average bacteria containing amount that requires each immobilized cell particle is 10
8Individual/particle, then with the Lu Shi yeast, give birth to fragrant Saite Candida and three kinds of immobilized cell particles of soy sauce tetrads by 2: 3: 0.5 the composite mixing of mass ratio.
(5) get the high-salt dilute soy stoste of 100kg step (3) gained, add the 10kg mass percent concentration and be 10% liquefaction syrup, the mixed bacteria immobilized cell particle for preparing is inoculated into soy sauce stoste, inoculum concentration is 30% (W
Mixed bacteria immobilized cell particle/ V
High-salt dilute soy stoste), ferment then, under 28 ℃ of fermentation temperatures, continuously fermented 24 hours, stir 2min every 30min, mixing speed 120r/min.
(6) after the fermentation ends, the soy sauce of fermenting-ripening is separated with the immobilized cell particulate filter, obtain high-salt dilute soy.Immobilized cell particle after the recovery can ferment next time.
Composition comparative analysis: utilize the method for SPME-gas chromatography mass spectrometry that the soy sauce sample of traditional handicraft fermentation in 120 days is analyzed with the volatile flavor component of the soy sauce sample that utilizes the immobilized cell technology fermentation.Testing result shows: utilize the content raising 18% of the soy sauce sample of immobilized cell fermentation than the 1-octene-3-alcohol in the soy sauce sample of fermentation in 120 days, 4-vinyl guajacolum content has improved 22.8%, and total ester has improved 9%, and ethyl acetate has improved 3%.
Sense organ compares: 120 days traditional handicraft fermenting-ripening soy sauce samples are carried out subjective appreciation with the soy sauce sample that utilizes the immobilized cell technology fermentation.Subjective appreciation is the result show: the soy sauce sample that utilizes the flavor of soy sauce of the soy sauce sample of immobilized cell technology fermentation to ferment than 120 days traditional handicrafts has improvement, and sauce perfume (or spice) is comparatively mellow.
Embodiment five:
Adopt immobilized cell fermentation to prepare the method for high-salt dilute soy:
(1) gets Lu Shi yeast, living fragrant Saite Candida respectively, be that the sucrose solution of 2% (W/W) activates under 30 ℃ of constant temperatures with mass percent concentration respectively, each barms vigor is fully recovered, spread cultivation then, centrifugal, stand-by with the sterilized water dilution, bacterial classification: sterilized water=1: 6 (W/V).
(2) soy sauce tetrads bacterial classification inoculation is activated more than 3 times in degreasing milk medium, spawn activity is fully recovered, it is centrifugal to spread cultivation then, stand-by with the sterilized water dilution, bacterial classification: sterilized water=1: 25 (W/V).
(3) the traditional high-salt dilute soy fermentation method of employing prepares the high-salt dilute soy stoste through fermenting 75 days, and is standby;
(4) preparation method of bacterial classification immobilization particle: with the Lu Shi yeast of step (1) and step (2) preparation, giving birth to fragrant Saite Candida and soy sauce tetrads bacterium liquid is that the sodium alginate glue of 1.5% (W/W) is mixed into cell suspending liquid by the volume ratio of 1: 23 (V/V) with mass percent concentration respectively, then cell suspending liquid being instilled into mass percent concentration is in 2% calcium chloride solution, wherein the volume ratio of 2% calcium chloride solution and sodium alginate cell suspending liquid is 2.5: 1 (V/V), promptly obtains the Lu Shi yeast respectively, give birth to the pearl immobilization calcium alginate particle of fragrant Saite Candida and soy sauce tetrads.The average bacteria containing amount that requires each immobilized cell particle is 10
7Individual/particle, then with the Lu Shi yeast, give birth to fragrant Saite Candida and three kinds of immobilized cell particles of soy sauce tetrads by 3: 4: 1 the composite mixing of mass ratio.
(5) get the high-salt dilute soy stoste of 100kg step (3) gained, add the 20kg mass percent concentration and be 5% liquefaction syrup, the mixed bacteria immobilized cell particle for preparing is inoculated into soy sauce stoste, inoculum concentration is 10% (W
Mixed bacteria immobilized cell particle/ V
High-salt dilute soy stoste), ferment then, under 32 ℃ of fermentation temperatures, continuously fermented 15 hours, stir 3min every 30min, mixing speed 100r/min.
(6) after the fermentation ends, the soy sauce of fermenting-ripening is separated with the immobilized cell particulate filter, obtain high-salt dilute soy.Immobilized cell particle after the recovery can ferment next time.
Composition comparative analysis: utilize the method for SPME-gas chromatography mass spectrometry that the soy sauce sample of traditional handicraft fermentation in 120 days is analyzed with the volatile flavor component of the soy sauce sample that utilizes the immobilized cell technology fermentation.Testing result shows: utilize the content raising 18.6% of the soy sauce sample of immobilized cell fermentation than the 1-octene-3-alcohol in the soy sauce sample of fermentation in 120 days, 4-vinyl guajacolum content has improved 20.7%, and total ester has improved 6%, and ethyl acetate has improved 2%.
Sense organ compares: 120 days traditional handicraft fermenting-ripening soy sauce samples are carried out subjective appreciation with the soy sauce sample that utilizes the immobilized cell technology fermentation.Subjective appreciation is the result show: the soy sauce sample that utilizes the flavor of soy sauce of the soy sauce sample of immobilized cell technology fermentation to ferment than 120 days traditional handicrafts has improvement, and sauce perfume (or spice) is comparatively mellow.
Embodiment six:
Adopt immobilized cell fermentation to prepare the method for high-salt dilute soy:
(1) gets torulopsis, living fragrant Saite Candida respectively, be that the sucrose solution of 2% (W/W) activates under 30 ℃ of constant temperatures with mass percent concentration respectively, each barms vigor is fully recovered, spread cultivation then, centrifugal, stand-by with the sterilized water dilution, bacterial classification: sterilized water=1: 6 (W/V).
(2) soy sauce tetrads bacterial classification inoculation is activated more than 3 times in degreasing milk medium, spawn activity is fully recovered, it is centrifugal to spread cultivation then, stand-by with the sterilized water dilution, bacterial classification: sterilized water=1: 25 (W/V).
(3) the traditional high-salt dilute soy fermentation method of employing prepares the high-salt dilute soy stoste through fermenting 90 days, and is standby;
(4) preparation method of bacterial classification immobilization particle: with the torulopsis of step (1) and step (2) preparation, giving birth to fragrant Saite Candida and soy sauce tetrads bacterium liquid is that the sodium alginate glue of 2.5% (W/W) is mixed into cell suspending liquid by the volume ratio of 1: 18 (V/V) with mass percent concentration respectively, then cell suspending liquid being instilled into mass percent concentration is in 2% calcium chloride solution, wherein the volume ratio of 2% calcium chloride solution and sodium alginate cell suspending liquid is 2: 1 (V/V), promptly obtains torulopsis respectively, give birth to the pearl immobilization calcium alginate particle of fragrant Saite Candida and soy sauce tetrads.Requiring the average bacteria containing amount of immobilized cell particle is 10
7Individual/particle, then with torulopsis, give birth to fragrant Saite Candida and three kinds of immobilized cell particles of soy sauce tetrads by 3: 3: 1 the composite mixing of mass ratio.
(5) get the high-salt dilute soy stoste of 100kg step (3) gained, add the 20kg mass percent concentration and be 5% liquefaction syrup, the mixed bacteria immobilized cell particle for preparing is inoculated into soy sauce stoste, inoculum concentration is 25% (W
Mixed bacteria immobilized cell particle/ V
High-salt dilute soy stoste), ferment then, under 30 ℃ of fermentation temperatures, continuously fermented 20 hours, stir 3min every 30min, mixing speed 100r/min.
(6) after the fermentation ends, the soy sauce of fermenting-ripening is separated with the immobilized cell particulate filter, obtain high-salt dilute soy.Immobilized cell particle after the recovery can ferment next time.
Composition comparative analysis: utilize the method for SPME-gas chromatography mass spectrometry that the soy sauce sample of traditional handicraft fermentation in 120 days is analyzed with the volatile flavor component of the soy sauce sample that utilizes the immobilized cell technology fermentation.Testing result shows: utilize the content raising 15% of the soy sauce sample of immobilized cell fermentation than the 1-octene-3-alcohol in the soy sauce sample of fermentation in 120 days, 4-vinyl guajacolum content has improved 13%, and total ester has improved 10%, and ethyl acetate has improved 5%.
Sense organ compares: 120 days traditional handicraft fermenting-ripening soy sauce samples are carried out subjective appreciation with the soy sauce sample that utilizes the immobilized cell technology fermentation.Subjective appreciation is the result show: the soy sauce sample that utilizes the flavor of soy sauce of the soy sauce sample of immobilized cell technology fermentation to ferment than 120 days traditional handicrafts has improvement, and sauce perfume (or spice) is comparatively mellow.
The present invention can summarize with other the concrete form without prejudice to spirit of the present invention or principal character.Above-mentioned embodiment of the present invention all can only be thought explanation of the present invention rather than restriction, therefore in implication suitable with claims of the present invention and any change in the scope, all should think to be included in the scope of claims.
Claims (8)
1. method that adopts immobilized cell fermentation to prepare high-salt dilute soy is characterized in that it may further comprise the steps:
(1) prepares soy sauce with traditional high-salt dilute fermentation method, make through the soy sauce stoste of fermentation in 60-90 days standby;
(2) learn from else's experience activation torulopsis, Lu Shi yeast, give birth to fragrant Saite Candida and four kinds of bacterial classifications of soy sauce tetrads are made the immobilized cell particle respectively;
The immobilized cell particle of (3) get the torulopsis that makes in the step (2), Lu Shi yeast, giving birth to fragrant Saite Candida and four kinds of bacterial classifications of soy sauce tetrads is according to mass ratio 0～1: carry out composite obtain mixed immobilization cell particle at 0～3: 3～5: 0.1～1; Be that 10-30% (W/V) is seeded to mixed immobilization cell particle in the high-salt dilute fermented sauce stoste of step (1) gained then with the inoculum concentration, continuously fermented 15-24 hour at temperature 28-33 ℃;
(4) after the fermentation ends, with after the high-salt dilute fermented sauce of this fermenting-ripening and the immobilized cell separate particles high-salt dilute fermented sauce finished product.
2. employing immobilized cell fermentation according to claim 1 prepares the method for high-salt dilute soy, it is characterized in that the immobilized cell particle of described torulopsis, Lu Shi yeast, living fragrant Saite Candida and four kinds of bacterial classifications of soy sauce tetrads makes by following steps:
The learn from else's experience torulopsis, Lu Shi yeast of activation, the bacterium liquid of giving birth to fragrant Saite Candida and four kinds of bacterial classifications of soy sauce tetrads is mixed with into cell suspending liquid with the sodium alginate glue by 1: 15～25 volume ratio respectively, then cell suspending liquid is instilled into mass percent concentration and is in 2% the calcium chloride solution, promptly obtain torulopsis, Lu Shi yeast respectively, give birth to the immobilized cell particle of fragrant Saite Candida and four kinds of bacterial classifications of soy sauce tetrads.
3. employing immobilized cell fermentation according to claim 1 and 2 prepares the method for high-salt dilute soy, it is characterized in that, the mass percent concentration of described sodium alginate glue is 1.5～3.5%.
4. employing immobilized cell fermentation according to claim 3 prepares the method for high-salt dilute soy, it is characterized in that, described mass percent concentration is that the addition of 2% calcium chloride solution and the volume ratio of sodium alginate bacterium liquid are 1.5～3: 1.
5. employing immobilized cell fermentation according to claim 4 prepares the method for high-salt dilute soy, it is characterized in that it is 10 that the immobilized cell particle of described torulopsis, Lu Shi yeast, living fragrant Saite Candida and four kinds of bacterial classifications of soy sauce tetrads all requires the average bacteria containing amount in the immobilized cell particle
6～10
8Individual/particle.
6. employing immobilized cell fermentation according to claim 5 prepares the method for high-salt dilute soy, it is characterized in that, the interpolation mass percent concentration is 5～10% liquefaction syrup in the high-salt dilute fermented sauce stoste of described step (2), and its addition is 5～20% of a high-salt dilute fermented sauce stoste gross weight.
7. employing immobilized cell fermentation according to claim 6 prepares the method for high-salt dilute soy, it is characterized in that, once stirs every 30～60min in the process of continuously fermenting described in the step (2).
8. employing immobilized cell fermentation according to claim 7 prepares the method for high-salt dilute soy, it is characterized in that, the time of described stirring is 2～5min, and the speed of stirring is in 80～150r/min scope.
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