BG61712B1 - Ferments for bulgarian yoghourt and method for their preparation - Google Patents

Abstract

The ferments can also be used for making other lactic acidtype products inluding such of non-lactic substrates. They consistof at least one strain of the group of Lactobacillus delbrueckiisubsp. Bulgaricus and/or at least one strain of the group ofStreptococcus salivarius subsp. Thermophilus which are authenticalcultures isolated in geographical regions of Bulgaria havingtraditionin making of Bulgarian yoghourt, in the form of mixed orsymbiotic, or monocultures. The invention also relates to a methodfor the preparation of the ferments by which monocultures andmixed or symbiotic ferments in DVS form are produced. The DVSforms are suitable for the preparation of production ferment andfor direct application for making Bulgarian yogourt underindustrial or residential conditions by other dairy products,inluding such of non-lactic subsrates.

Description

Technical field

The invention relates to ferments intended for the production of Bulgarian yoghurt and lactic acid products, including non-dairy substrates, and to methods for their preparation.

BACKGROUND OF THE INVENTION

Symbiotic yeast is known for Bulgarian yogurt (1), consisting of the strain Lactobacillus bulgaricus selected from the group of strains Lactobacillus bulgaricus No. 5, 26, 37 or 144, and the strain Streptococcus thermophilus selected from the group Streptococcus thermophilus No. 14 , 17 and 22, in a ratio of 1: 3 to 1:10. Specifically, yeasts described are described, consisting of: L.b.N 5 + S.t.N 12, L.b.N 5 S.t.N 22, L.b.N 37 + S.t.N 18, L.b.N 37 + S.t.N 12, L.b.N 144 + S.t.N 12, L.b.N 144 + S.t.N 14, L.b. 26 + S.t.12.

The method of producing these yeasts consists of inoculating sterile cow's milk with various combinations of selected strains of Lactobacillus bulgaricus and Streptococcus thermophilus, including the above strains, separating combinations with normal morphology and normal ratios, which are mixed in sterile or in a mixture of sterile sheep's and cow's milk is a follow-up for 6 months, followed by combinations having stable morphological properties and a stable ratio of lactobacilli cell counts to streptococcus eyelets are used to prepare production yeasts. The inoculation and cultivation were carried out at 45 ° C ± 1 ° C. The cultivation was carried out for 2 h. Sowing is carried out daily for the first 4 months and once a week for the next two months, with the first sowing being carried out in a mixture of sheep's and cow's milk in a 1: 1 ratio.

A symbiotic yeast (2) known to be comprised of one or more strains of Lactobacillus delbrueckii subsp. bulgaricus NN by NBPCC 2183, 2184, 2185, 2186, 2187, 2188,

2189, 2190, 2191, 2192 and strains of Streptococcus salivarius subsp.thermophilus NN 2173, 2174, 2175, 2176, 2177, 2178, 2179, 2180, 2181, 2182. According to (2), concentrated yeast is produced as combinations of the above strains are cultured on nutrient media containing 1 part filtered fermented casein hydrolyzate and 2 parts by-products, with prior neutralization of the medium to pH 6.5-7.0 and sterilization at 120 ° C for 20 min at 45 ° ± 1 ° C, periodic neutralization in the range of pH 5.0 to 6.5, addition of gelling agent in the amount of 0.05 to 0.2% and lyophilization. Said secondary dairy product is a mixture of permeate or nail with digested lactose and / or sterile skimmed milk.

SUMMARY OF THE INVENTION

Yeast for the production of Bulgarian yogurt can also be used as a yeast for the production of other lactic acid products, including those from non-dairy substrates consisting of at least one strain selected from the group Lactobacillus delbrueckii subsp.bulgaricus, registered with NBPCC on 18.05. 1985 under number LBB B 5-273; LBB B26-285; LBB B 37-286; LBB B 144-287 and 14.12.1994 under the numbers: LBB B ml-2471; LBB B 31-2481; LBB B m2-2472; LBB B ml4-2473; LBB B 41-2482; LBB B 120-2483; LBB B 142478; LBB B 122-2484; LBB B 24-2479; LBB B 129-2485; LBB B 282-2480; LBB B 156-2486 and / or at least one strain selected from the group: Streptococcus salivarius subsp. thermophilus, registered with NBPMCC on 18.05.1985 under the numbers: LBB T 12-450; LBB T 14-451; LBB T 18-452; LBB T 22-453; and on 14 December 1994 under number LBB T 3-2487; LBB T 8-2488; LBB T 10-2489; LBB T 28-2492; LBB T 36-2493; LBB T 38-2494; LBB T 16-2490; LBB T t5-2474; LBB T 17-2491; LBB T R6-2475 as a mixed or symbiotic culture or monoculture.

The strains are authentic cultures isolated from geographical regions with traditional Bulgarian yogurt production and differentiated according to their morphological, physiological and biochemical characterization characteristics according to the requirements of the Birgi 1986 identifier and in IDF Standard 146/1991 and IDF Standard 149/1991.

The biochemical properties of the above strains are characterized by carbohydrate fermentation data and acidity and volatile aroma content.

Fermentation data of carbohydrates of Lactobacillus bulgaricus strains

Name of [ the strains ι Growth in the presence of ! glucose fructose lactose! saccharose ___________________________________________________1 I others * LBB In ml I + + + 1 LBB B m2 J + + ¹ LBB B m14: + + LBB B 5 + + . ..... + __________: ____________ LBB B 14; + + + -, - LBB B 24 I + + + - - LBB B 26 ^р + + + J-. . ..... LBB B 28! + + + - ι - . _{LB0 B 31} + + + +: 1 LBB B 37 j + + + i LBB B 41 | + · + + j- LBB B 120 i + + + J _________-_________ 1 ____-____ LBB B12? + ΐ + ¹ LBB B129 f + + + ί LBB B 144! + + + 1 LBB B 156 + ⁺ + ΐ

* - Glycerin, erythritol, D arabinose, L arabinose, ribose, D xylose, adonite, β-methyl-xylose, galactose, D mannose, L sorbose, rhamnose, dulcite, inositol, mannitol, sorbitol, α-methyl-0-mannose , os-methyl-D glucoside, N acetyl-glucosamone, amygdalin, arbutin, esculin, salicin, cellobiose, maltose, melibiosis, trehalose, inulin, melitisitose, D raffinose, amidon, glycogen, xylitol, β-gentianose D / ixose, D-tagatose, D-fucose, L-fucose, D-arabite, gluconate, 2 cetogluconate, 5 ceto-gluconate

Legend: / + / - fermented / - / - unfermented

Fermentation data for carbohydrates of strains

Streptococcus therm ophilus

Name of the strains Growth in the presence of glucose fructose lactose saccharose others * LBBT3 + + + + LBBT8 + + + + LBBT10 + + + + LBB T 12 +. + + + LBB T 14 + + + + LBB T 16 + + + + . LBB T 17 + + - ϊ D - LBB T 18 + + + - LBBT22 + + + + - LBB T 28 + + + + LBBT36 + + + + LBBT38 + + + + - LBBTm5 + + + + - LBB T R6 + + ± J + -

• - L arabinose, D arabinose, glycerin, inulin, maltose, mannitol, raffinose, rhamnose, ribose, oalicin, oorbitol, xylose, trehalose

Legend: / + / - fermented / - / - unfermented

Characteristics of certain strains of Lactobacillus b1 and dapsizun6 \ lrani in sterile whole cow's milk n = 3

Name to the strain D geographical origin Thermostat time / b / Acidity Volatile aromatic substances milky D-ml. Acetal pH to to dehyd Ethanol % % ppm ΡΡΠΐ LBB.B5 Deep 6 4.66 0.54 0 34 9 711 3.001 336 3.67 1.76 1.30 1.522 2435 LBB B26 Dissection 7 4.64 0.63 0.44 5 622 2.839 336 3.59 1.05 1.34 1.293 2,341 LBB.B28 Graschitsa 6 4.72 0.56 0.35 9.056 3,349 336 3.67 1.53 1.19 2.510 2,642 LBB.B37 Baptism 6 4.51 0.52 0.30 7.859 3,083 336 3.50 1.90 1.52 1.301 2.640 LBB.B41 Baptism 6 4.62 0.50 D 0.34 Ϊ0.788 2.232 336 3.56 1.62 1.30 1.085 2.142 LBB.B122 Archar - 6 4.61 0.62 0.42 4.30 3.09 Vidin 336 3.60 1.60 1.25 0.78 2.30 LBB.B144 New village 6 4.67 0.54 0.31 5.15 2.22 336 3.60 1.60 1.18 0.87 1.42

D pH - 0.15

Δ ml. to% - 0.1

6D / - / - 0.1

Characteristics of some strains of Streptococcus thermophilus incubated in sterile whole cow's milk n ~ 3

Name to the strain Geographical origin Thermostat time / AND / Acidity Volatile aromatic substances pH milk to% D ml. to% Acetal dehydrate ppm Ethanol PPt St. 13 Drain 6.30 336 4.65 3.92 0.60 1.08 0.45 0 92 1.23 0.54 1.40 1.41 St. t 8 Drain 5.30 336 4.66 4.07 0.59 1.04 0.41 0.84 2.62 0.91 1.70 1.44 st. t io Drain 5.30 336 4.68 4.08 0.61 1.08 0.42 0 85 2.82 0.93 1.93 1.48 St. 112 Graschitsa 6.30 467 0.59 0.41 2 93 208 336 3.89 1.08 0.94 0.71 1.77 St. t 14 Deep 5.30 336 4.65 3.89 0.58 1.04 0.41 0.79 2.24 0 70 1. 74 1.56 St. t 16 Deep 6.30 336 4.71 4.06 0.57 1.08 0.41 0.96 2.01 0.55 1.97 0.94 St. t 17 Hollows 5.30 336 4.71 4.06 0.58 1 06 0.41 0.80 1.95 0.75 0.85 0.70 St 118 Hollows 6.30 4.64 0.61 0.44 2.27 2 42 336 4.00 0.01 0.84 0.84 1.58 St. t 22 Famous 6.30 4.64 0.59 0.40 2.26 1.58 336 3.88 0.99 0 93 0.71 2.21 • St. t 38 Radnevo 7 4.60 0.63 0 39 1 86 2.76 _ 336 4.16 0.98 0.77 0 59 I.9G

Δ pH - 0.15

Δ ml. to% - 0.1

ΔΟ / - / - 01

Preferred variants of the symbiotic yeast according to the invention are combinations of Lactobacillus delbrueckii subsp.bulgaricus and Streptococcus salivarium subsp. thermophilus c numbers from NBPMK.K as follows: 2484 + 2491; 2482 + 451; 2484 + 453; 2480 + 453; 2478 + 2489; 2479 + 2488; 2479 + 2490; 2485 + 2490; 2486 + 453.

The method for producing mixed yeasts consists of inoculating monocultures from said strains, selected by colony morphology in a liquid nutrient medium with a dry matter content of 5 to 10%, consisting of skimmed milk, whose casein and lactose are hydrolyzed, and biologically an activating additive comprising a mesopeptone broth containing a meat extract, preferably 1-2%, peptone, preferably 0.5-1%, yeast extract, preferably 0, ΙΟ, 5% and cultivation at 37-42 ° C to pH 4.6 ± 0.1.

The monocultures thus obtained from

Lactobacillus delbrueckii subsp.bulgaricus and Streptococcus salivarius subsp.thermophilus can be used either as monocultures or to produce mixed starter cultures.

The method for producing symbiotic yeast consists in maintaining, by repeated transplantation, for at least 4 months, combinations of strains of Lactobacillus delbrueckii subsp.bulgaricus and Streptococcus salivarius subsp.thermophilus, which, when co-cultured in skimmed or whole foods or in whole fat or whole foods 1 have shown stable symbiotic relationships within the desired range.

The method of producing a monoculture concentrate or concentrated mixed or symbiotic yeast for direct application (DVS) consists of inoculating with 3 to 8% of the respective culture medium II, consisting of reconstituted dry whey and buffer-activating mixture representing an aqueous solution of : CH ₃ COONa, preferably from 5 to 10%, KN ₂ PO ₄ , preferably 1-4%, Na ₂ HPO ₄ , preferably 1-4%, MgSO ₄ , preferably 0.1-0.5% , MnSO ₄ , preferably 0.01-0.06%, yeast extract, preferably 0.1-0.5%; or in culture medium III consisting of reconstituted skimmed milk powder and a buffer-activating mixture representing an aqueous solution of: CH ₃ COONa, preferably from 5 to 10%, KN ₂ PO ₄ , preferably 1-4%, Na ₂ HPO ₄ , preferably 1-4%, MgSO ₄ , preferably 0.1-0.5%, MnSO ₄ , preferably 0.01-0.06%, yeast extract, preferably 0.1-0.5%, and lactose and protein concentrate obtained by ultrafiltration, preferably 0.5-4%, the fermentation process in culture medium II or III lasting 4-5 hours, after which the resulting culture is mixed with n rotation medium and equilibration is carried out at 15 to 20 ° C.

The protection medium is a sterile aqueous solution of sucrose, sodium glutamate and vitamin C or polyethylene oxide (PE0) - mol. 400 or 1000. The concentrated culture thus prepared is frozen at -65 to -80 ° C or -196 ° C or lyophilized.

The advantages of the strains according to the invention are that they allow to obtain yeasts representing monocultures or mixed and symbiotic cultures of different combinations of at least one strain selected from the group Lactobacillus delbrueckii subsp.bulgaricus according to the invention and at least one strain selected from the group of Streptococcus salivarius subsp.thermophilus according to the invention in desired proportions, yielding essentially the same lactic acid product, but with known differences in its properties according to different taste preferences or specific conditions. st production and storage.

Another advantage of the strains according to the invention is that the symbiotic starters obtained with them retain the desired ratio between Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus salivarius subsp. thermophilus. Quality standardized yeast is obtained.

The method and the nutrient media according to the invention make it possible to obtain monocultures, mixtures or symbiotic starters such as DVSform, whereby the process is significantly shortened compared to the known method of producing concentrated starters.

The resulting DVS cultures (mono- or mixed or symbiotic) are suitable for the production of yeast production and for direct application for the production of Bulgarian yogurt in industrial and domestic conditions, as well as for the production of other lactic acid products, including those from non-dairy substrates, which makes it possible to produce various dietary and prophylactic foods.

Embodiments of the invention

Example 1. Preparation of monocultures and mixed starters.

Pure cultures of Lactobacillus delbrueckii subsp.bulgaricus and Streptococcus salivarius subsp.thermophilus incubated for 16 h at 37 ° C in liquid culture medium (MRS or Ml7) were seeded in solid culture medium (MRS or Ml7 / in the amount required to obtain single colonies incubated for 48 h at 37 ° C. Colonies with characteristic morphology were selected and plated in liquid culture medium. Incubated for 18 h at 37 ° C. With 5 1 of the resulting laboratory culture, culture medium I was inoculated. 100 1 skimmed milk, whose casein and lactose are hydrolyzed, meat extract 100 g, peptone 50 g, yeast extract 10 g and water to a solids content of 8%, neutralized with ammonia to pH 6,5 and incubated at 42 ° C until pH 4,6 is achieved. The number of microorganisms is reached from 10 'to 10 ^s cfu / ml for strains of Lactobacillus delbrueckii subsp.bulgaricus and 10 ^s to 10 'cfu / ml for strains of Streptococcus salivarius subsp.thermophilus.

The monocultures thus obtained from the said strains of Lactobacillus delbrueckii subsp.bulgaricus and Streptococcus salivarius subsp.thermophilus are used to prepare mixed yeasts for Bulgarian yogurt, combined in a ratio of 1: 1 to 1:15.

Example 2. Preparation of concentrated yeast.

100 1 recovered dry whey (6% dry matter) was mixed with 2.5 1 buffer-activating mixture representing aqueous solution of CH ₃ COONa - 10%, CN ₂ PO ₄ - 4%, Na ₂ HPO ₄ 4%, MgSO ₄ - 0.2%, MnSO ₄ - 0.04% and yeast extract - 0.2%. Thermize at 115 ° C for 10 min.

The medium was cooled to 45 ° C and inoculated with 5 I refreshed laboratory symbiotic yeast consisting of Lactobacillus delbrueckii subsp.bulgaricus per. No. NBPMC 2480 and Streptococcus salivarius subsp. thermophilus NBMPCC N ° 453. Incubate in a fermentor at 43 ° C for 45 h at pH 5.5-6.0 under automatic pH and temperature control. The resulting concentrated yeast was cooled to 4 ° C, mixed with 10 l of a protective medium representing a sterile aqueous solution of 10% sucrose, 2% sodium glutamate and 0.1% vitamin C and lyophilized.

In another embodiment, the protective medium is polyethylene oxide in an amount of 1% aqueous solution of PEO-400 or PEO-0000.

Example 3. Preparation of concentrated yeast.

100 1 recovered skimmed milk powder (6% dry matter) is mixed with 2.5 1 buffer-activating mixture representing an aqueous solution of 4% lactose, 1% protein concentrate, 10% CH ₃ COONa, 4% CN ₂ PO ₄ , 4% Na ₂ HPO ₄ , 0.2% MgSO ₄ , 0.4% MnSO ₄ and 0.2% yeast extract were thermalized at 115 ° C for 10 min. Cool to 45 ° C and inoculate with 5 l of laboratory symbiotic yeast consisting of Lactobacillus delbrueckii subsp.bulgaricus per. No. NBPMCC 2482 and Streptococcus salivarius subsp.thermophilus NBPMCC No. 451 or with other symbiotic cultures of the invention. The cultivation and preparation of the concentrated symbiotic culture was continued as described in Example 2.

Example 4. Preparation of concentrated yeast.

100 l of nutrient medium consisting of reconstituted dry whey and 2.5% buffer-activating mixture as in Example 3 was treated according to the procedure described in Example 2. It is inoculated with 5 l of the refreshed monoculture Streptococcus salivarius subsp.thermophilus NBPMCC No. 2491. The incubation is continued for 6-7 h at 42 ° C and pH 5.7-6.0 under automatic pH and temperature control. The concentrated and cooled monoculture was mixed with 10 l of the protection medium described in Example 2 and lyophilized.

Example 5. Preparation of concentrated monoculture.

100 l of the culture medium as in Examples 2 and 3 and treated according to the procedure described in Example 2 were inoculated with 5 l of the refreshed monoculture from Lactobacillus delbrueckii subsp.bulgaricus per. No. NBPMC 2479. The procedure was continued analogously to Example 4.

Example 6. Mixed DVS culture.

Lyophilized concentrated culture of Lactobacillus delbrueckii subsp.bulgaricus per. No. NBPMCC 2484 prepared according to Example 5 and Streptococcus salivarius subsp.thermophilus NBPMCC No. 2491 prepared according to Example 4 were mixed in a ratio of 1: 1 to 1: 5 and used as DVS leaven.

By analogy to the above examples, other strains of the two species according to the invention are obtained monocultures or mixed, incl. multilayered or symbiotic yeasts in liquid, frozen or lyophilized form.

According to the invention, the original Bulgarian yoghurt can be made from fresh or reconstituted dry sheep, cows, buffalo and other milk, as well as a wide range of lactic acid products, including non-dairy substrates such as soybean flour, oatmeal, etc.

DVS yeasts can be used to obtain production yeasts or for direct application for the production of Bulgarian yoghurt and other lactic acid products.

Claims (8)

1. Yeast for Bulgarian yoghurt and lactic acid products containing strains of the species Lactobacillus bulgaricus and Streptococcus thermophilus, characterized in that it consists of at least one strain selected from the group Lactobacillus delbrueckii subsp.bulgaricus registered in NB.05.05M85 d. under LBB numbers B 5-273; LBB B 26-285; LBB B 37286; LBB B 144-287 and 14.12.1994 under the numbers: LBB B ml-2471; LBB B 31-2481; LBB B m2-2472; LBB B ml4-2473; LBB B 41-2482; LBB B 120-2483; LBB B 14-2478; LBB B 122 2484; LBB B 24-2479; LBB B 129-2485; LBB B 282-2480; LBB B 156-2486 and / or at least one strain selected from the group: Streptococcus salivarius subsp. thermophilus, registered with NBPMCC on 18.05.1985 under the numbers: LBB T 12-450; LBB T 14-451; LBB T 18-452; LBB T 22-453; and on 14.12.1994 under the numbers: LBB T 3-2487; LBB T 8-2488; LBB T 10-2489; LBB T 28-2492; LBB T 36-2493; LBB T 38-2494; LBB T 16-2490; LBB T P15-2474; LBB T 17-2491; LBB T R6-2475 as a mixed or symbiotic culture or monoculture. 2. A method for producing monocultures or mixed starters, comprising the cultivation of monocultures of the strains of claim 1 constituting authentic cultures isolated from Bulgarian yogurt, characterized in that the selected monocultures are sown in a liquid culture medium I consisting of skimmed milk, the casein and lactose of which have been hydrolyzed, a mesopeptone broth, water and a biologically activating additive selected according to the type of lactic acid microorganism, followed by cultivation at 42 ° C to pH 4,6 + 0,1. A method for producing monocultures or mixed starters according to claim 2, characterized in that the sowing is carried out in nutrient medium I with a dry matter of 5 to 10%, consisting of skimmed milk, whose casein and lactose are hydrolyzed, and a biologically activating additive consisting of a mesopeptone broth containing a meat extract, preferably 12%, a peptone, preferably 0.5-1%, a yeast extract, preferably 0.1-0.5%. 4. A method of producing symbiotic yeast, characterized in that they are selected by repeated transplantation into a culture medium 1 consisting of skimmed milk, whose casein and lactose have been hydrolyzed, a mesopeptone broth, water and a biologically activating additive selected by the species lactic acid microorganism, for at least 4 months, of combinations of strains of Lactobacillus delbrueckii subsp.bulgaricus and Streptococcus salivarius subsp.thermophilus, as claimed in claim 1, in a ratio of 1: 1 to 1: 5. 5. A method of producing a concentrated mixed or symbiotic yeast or monoculture for direct administration, characterized in that it consists of inoculation with the respective culture medium II, consisting of reconstituted dry whey and a buffer-activating mixture comprising an aqueous solution of CH ₃ COONa, KN ₂ PO ₄ , Na ₂ HPO ₄ , MgSO ₄ , MaSO ₄ and yeast extract, cultured in fermenter at 43-45 ° C for 4-5 h at pH 5.5-6, cooled to 4 -6 ° C, after which the resulting concentrated culture was mixed with a protection medium consisting of sorghum, sodium glutamate and vitamin C or polyethylene oxide - PEO-400 or PEO-1000 and frozen at -65 to 80 ° C or at -196 ° C or lyophilized. A method of producing concentrated mixed or symbiotic yeast or monoculture according to claim 5, characterized in that the respective culture is inoculated into culture medium II, consisting of reconstituted dry whey and a buffer-activating mixture comprising an aqueous solution of: CH ₃ COONa 5 to 10%, CN ₂ ₽ ₄ - 14%, Na ₂ HPO ₄ - 1-4%, MgSO ₄ - 0.1-0.5%, MnSO ₄ - 0.01-0.06%, yeast extract - 0,10,5%. 7. A method of producing concentrated mixed or symbiotic yeast or monoculture for direct administration, characterized in that it consists of inoculation with the respective culture medium III, consisting of reconstituted skimmed milk powder and a buffer-activating mixture comprising an aqueous solution of lactose and protein concentrate obtained by ultrafiltration, CH ₃ COONa, CN ₂ PO ₄ , Na ₂ HPO ₄ , MgSO ₄ , MaSO ₄ and yeast extract, cultured in fermenter at 43-45 ° C for 4-5 h at pH 5.5-6, cooling to 4-6 ° C, then the resulting horse centered culture is mixed with a protective medium consisting of sucrose, sodium glutamate and vitamin C or polyethylene oxide - PEO-400 or PEO-1000 and frozen at -65 to 80 ° C or -196 ° C or lyophilized. A method for producing concentrated mixed or symbiotic yeast or monoculture according to claim 7, characterized in that the respective culture is inoculated into culture medium III, consisting of reconstituted skimmed milk powder and a buffer-activating mixture representing an aqueous solution of: CH ₃ COONa - 5-10%, KN ₂ PO ₄ 1-4%, Na ₂ HPO ₄ - 1-4%, MgSO ₄ - 0.1-0.5%, MnSO ₄ - 0.01-0.06%, yeast extract - 0, ΙΟ, 5% and lactose and protein concentrate obtained by ultrafiltration - 0,5-4%.