JPS63202390A - Method for enhancing amount of produced diacetyl by lactic acid bacteria - Google Patents
Method for enhancing amount of produced diacetyl by lactic acid bacteriaInfo
- Publication number
- JPS63202390A JPS63202390A JP3337887A JP3337887A JPS63202390A JP S63202390 A JPS63202390 A JP S63202390A JP 3337887 A JP3337887 A JP 3337887A JP 3337887 A JP3337887 A JP 3337887A JP S63202390 A JPS63202390 A JP S63202390A
- Authority
- JP
- Japan
- Prior art keywords
- lactic acid
- diacetyl
- acid bacteria
- citric acid
- amount
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 title claims abstract description 46
- QSJXEFYPDANLFS-UHFFFAOYSA-N Diacetyl Chemical group CC(=O)C(C)=O QSJXEFYPDANLFS-UHFFFAOYSA-N 0.000 title claims abstract description 41
- 241000894006 Bacteria Species 0.000 title claims abstract description 24
- 239000004310 lactic acid Substances 0.000 title claims abstract description 23
- 235000014655 lactic acid Nutrition 0.000 title claims abstract description 23
- 238000000034 method Methods 0.000 title claims description 8
- 230000002708 enhancing effect Effects 0.000 title claims description 6
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 claims abstract description 96
- 229910052742 iron Inorganic materials 0.000 claims abstract description 9
- 229910052750 molybdenum Inorganic materials 0.000 claims abstract description 9
- 150000003839 salts Chemical class 0.000 claims abstract description 9
- 229910052802 copper Inorganic materials 0.000 claims abstract description 7
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 claims description 15
- 238000000855 fermentation Methods 0.000 claims description 11
- 230000004151 fermentation Effects 0.000 claims description 11
- 239000000758 substrate Substances 0.000 claims description 11
- 239000010949 copper Substances 0.000 claims description 7
- 239000011733 molybdenum Substances 0.000 claims description 7
- 230000001965 increasing effect Effects 0.000 claims description 6
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 claims description 5
- ZOKXTWBITQBERF-UHFFFAOYSA-N Molybdenum Chemical compound [Mo] ZOKXTWBITQBERF-UHFFFAOYSA-N 0.000 claims description 5
- -1 iron ions Chemical class 0.000 claims description 4
- JPVYNHNXODAKFH-UHFFFAOYSA-N Cu2+ Chemical compound [Cu+2] JPVYNHNXODAKFH-UHFFFAOYSA-N 0.000 claims description 2
- 229910001431 copper ion Inorganic materials 0.000 claims description 2
- 241000192129 Leuconostoc lactis Species 0.000 claims 1
- 235000020183 skimmed milk Nutrition 0.000 abstract description 6
- 239000001963 growth medium Substances 0.000 abstract description 4
- 239000000843 powder Substances 0.000 abstract description 4
- 235000021001 fermented dairy product Nutrition 0.000 abstract description 3
- 125000003118 aryl group Chemical group 0.000 abstract description 2
- 239000004615 ingredient Substances 0.000 abstract description 2
- 239000007787 solid Substances 0.000 abstract description 2
- 150000002500 ions Chemical class 0.000 abstract 3
- 241000194035 Lactococcus lactis Species 0.000 abstract 1
- 235000014897 Streptococcus lactis Nutrition 0.000 abstract 1
- 230000000694 effects Effects 0.000 description 13
- 238000004519 manufacturing process Methods 0.000 description 10
- 241000194017 Streptococcus Species 0.000 description 9
- 108090000790 Enzymes Proteins 0.000 description 7
- 102000004190 Enzymes Human genes 0.000 description 7
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 6
- 239000002609 medium Substances 0.000 description 6
- 230000001580 bacterial effect Effects 0.000 description 5
- 108010036824 Citrate (pro-3S)-lyase Proteins 0.000 description 4
- 238000000691 measurement method Methods 0.000 description 4
- 238000012258 culturing Methods 0.000 description 3
- 235000013365 dairy product Nutrition 0.000 description 3
- 230000004060 metabolic process Effects 0.000 description 3
- 239000008363 phosphate buffer Substances 0.000 description 3
- 239000001509 sodium citrate Substances 0.000 description 3
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 3
- 244000172809 Leuconostoc cremoris Species 0.000 description 2
- 235000017632 Leuconostoc cremoris Nutrition 0.000 description 2
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L Magnesium chloride Chemical compound [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 description 2
- 235000008429 bread Nutrition 0.000 description 2
- 235000014121 butter Nutrition 0.000 description 2
- 235000013310 margarine Nutrition 0.000 description 2
- 239000003264 margarine Substances 0.000 description 2
- 229910021645 metal ion Inorganic materials 0.000 description 2
- 235000013336 milk Nutrition 0.000 description 2
- 239000008267 milk Substances 0.000 description 2
- 210000004080 milk Anatomy 0.000 description 2
- 239000011148 porous material Substances 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 239000008223 sterile water Substances 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 235000007294 Brassica nipposinica Nutrition 0.000 description 1
- 241000342995 Brassica rapa subsp. nipposinica Species 0.000 description 1
- KRKNYBCHXYNGOX-UHFFFAOYSA-K Citrate Chemical compound [O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O KRKNYBCHXYNGOX-UHFFFAOYSA-K 0.000 description 1
- 229910021578 Iron(III) chloride Inorganic materials 0.000 description 1
- 241000634997 Leucon Species 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 239000012190 activator Substances 0.000 description 1
- 238000009835 boiling Methods 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 235000013351 cheese Nutrition 0.000 description 1
- 230000015271 coagulation Effects 0.000 description 1
- 238000005345 coagulation Methods 0.000 description 1
- 235000016213 coffee Nutrition 0.000 description 1
- 235000013353 coffee beverage Nutrition 0.000 description 1
- 238000004040 coloring Methods 0.000 description 1
- 235000015140 cultured milk Nutrition 0.000 description 1
- 235000015142 cultured sour cream Nutrition 0.000 description 1
- 239000000706 filtrate Substances 0.000 description 1
- 235000012907 honey Nutrition 0.000 description 1
- UQVDQSWZQXDUJB-UHFFFAOYSA-N hydron;7h-purin-6-amine;chloride Chemical compound Cl.NC1=NC=NC2=C1NC=N2 UQVDQSWZQXDUJB-UHFFFAOYSA-N 0.000 description 1
- 229910017053 inorganic salt Inorganic materials 0.000 description 1
- RBTARNINKXHZNM-UHFFFAOYSA-K iron trichloride Chemical compound Cl[Fe](Cl)Cl RBTARNINKXHZNM-UHFFFAOYSA-K 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 229910001629 magnesium chloride Inorganic materials 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 229940037201 oris Drugs 0.000 description 1
- 230000037361 pathway Effects 0.000 description 1
- 239000002243 precursor Substances 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 235000013580 sausages Nutrition 0.000 description 1
- 239000007858 starting material Substances 0.000 description 1
- 238000002525 ultrasonication Methods 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
- 235000013618 yogurt Nutrition 0.000 description 1
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
Description
【発明の詳細な説明】
〔産業上の利用分野〕
本発明は、乳酸菌の産生ずる主要な香気成分であるジア
セチルの生成量の増強法に関するものである。DETAILED DESCRIPTION OF THE INVENTION [Industrial Application Field] The present invention relates to a method for enhancing the amount of diacetyl produced, which is a major aroma component produced by lactic acid bacteria.
ジアセチルは醗酵バター、サワークリーム、チーズなど
の醗酵乳製品の主要な香気成分であり、マーガリン、サ
マーソーセージ、はち蜜、コーヒー、パンなどでは芳香
として喜ばれている。乳酸菌によるジアセチル生成能あ
るいは乳製品中のジアセチル生成量を高めるためにジア
セチルの生成前駆体であるクエン酸を添加して培養する
方法が古くから報告されており、またＬｅｕｃｏｎｏｓ
ｔｏｃ ｃｒｅ−ｍｏｒｉｓにおいてはＭｎ”の添加が
水菜の増殖ならびにクエン酸の利用性を高めることが報
告されている（Ｔ、Ｍ、Ｃｏｇａｎ、Ｊ、Ｄａｉｒｙ
Ｒｅ５ｅａｒｃｈ、４２．１３９（１９７５））。Diacetyl is a major aroma component in fermented dairy products such as fermented butter, sour cream, and cheese, and is also a welcome aromatic component in margarine, summer sausage, honey, coffee, bread, and other products. In order to increase the diacetyl production ability of lactic acid bacteria or the amount of diacetyl produced in dairy products, a method of culturing with the addition of citric acid, which is a production precursor of diacetyl, has been reported for a long time.
It has been reported that the addition of Mn'' to M. toc cre-moris increases the growth of mizuna and the availability of citric acid (T, M, Cogan, J, Dairy
Re5arch, 42.139 (1975)).
しかしＳｔｒｅｐｔｏｃｏｃｃｕｓ １ａｃｔｉｓ ５
ｕｂｓｐ、 ｄｉａｃｅｔｙｌａ−ｃｔｉｓにおいては
、培地中へのクエン酸の添加を除いてジアセチルの生成
を促進する有効な成分は認められていない。However, Streptococcus 1actis 5
In the ubsp, diacetyla-ctis, no effective ingredient has been found to promote diacetyl production except for the addition of citric acid to the medium.
また、乳酸菌によるジアセチル生成機構については、こ
れまでＲ，Ａ、Ｓｐｅｃｋｍａｎら（Ｊ、Ｂａｃｔ、、
９５，１７４（１９６８））＋ Ｊ、Ｓｔａｄｈｏｕ
ｄｅｒｓ（Ｍｉｌｃｈｗｉｓｓｅｎｓｃｈａｆｔ＋ ２
９＋３２９ （１９７４））等の報告がある。いずれも
クエン酸を代謝してジアセチルを生成する経路につき記
載している。Furthermore, regarding the diacetyl production mechanism by lactic acid bacteria, R.A., Speckman et al.
95, 174 (1968)) + J, Stadhou
ders (Milchwissenschaft + 2
There are reports such as 9+329 (1974)). All of them describe the pathway that metabolizes citric acid to produce diacetyl.
さらにＳｔｒｅｐｔｏｃｏｃｃｕｓ Ｉａｃｔｔｓ ５
ｕｂｓｐ、 ｄｉａｃｅｔｙｌ−ａｃｔｉｓによるジア
セチルの生成量を高めるために紫外線照射による変異株
の使用（Ｒ１に、Ｋｕｉｌａｊ、Ｄａｔ−ｒｙ Ｓｃｉ
、、６１．３７９（１９７８））、固定化菌体の使用（
Ｊ。Furthermore, Streptococcus Iactts 5
Use of mutant strains by ultraviolet irradiation to increase the amount of diacetyl produced by ubsp, diacetyl-actis (R1, Kuilaj, Dat-ry Sci.
, 61.379 (1978)), use of immobilized bacterial cells (
J.
Ｒｏｓｓ ｉ、 Ｍｉ ｌｃｈｗｉｓｓｅｎｓｃｈａｆ
ｔ、 ３９＋ ３３６ （１９８４） ）などが報告
されている。Rossi, Milchwissenschaf
T, 39+336 (1984)), etc. have been reported.
乳酸菌によるジアセチル生成量を高めるための上記の従
来技術につぎのような改良を加えた。すなわち培地中の
クエン酸を効率よくジアセチルに変換させるための方法
につき種々検討を重ねたものである。The following improvements were made to the above conventional technology for increasing the amount of diacetyl produced by lactic acid bacteria. That is, various studies have been conducted on methods for efficiently converting citric acid in a culture medium into diacetyl.
本発明は、ジアセチル生成促進物質を培養基質に添加す
るだけの簡単な手段により、ジアセチル生成量を高める
ことを目的とするものである。The object of the present invention is to increase the amount of diacetyl produced by simply adding a diacetyl production promoting substance to the culture substrate.
本発明者らはＳｔｒｅｐｔｏｃｏｃｃｕｓ １ａｃｔｉ
ｓ ５ｕｂｓｐ、ｄｉ−ａｃｅｔｙｌａｃｔｉｓ、Ｌｅ
ｕｃｏｎｏｓｔｏｃ ｃｒｅｍｏｒｉｓ＋Ｌｅｕｃｏｎ
ｏｓｔｏｃＩａｃｔｉｓ等のジアセチル生成に関する諸
酵素活性の活性化因子につき鋭意研究を重ねた結果、ジ
アセチル生成促進のためにはクエン酸とともにＣｕ２“
。The present inventors have discovered that Streptococcus 1acti
s 5ubsp, di-acetylactis, Le
uconostoc cremoris+Leucon
As a result of intensive research on the activators of various enzyme activities related to diacetyl production such as S. ostocIactis, we found that Cu2" along with citric acid is necessary to promote diacetyl production.
.
ｐ ｅ ２ ＋ 、 Ｍ Ｏｈ ＋ 、 Ｆｅ ３ ＋
の１種または２種以上を含有するようにこれらの塩類を
それぞれ強化した培地で培養するとジアセチルの生成量
を増大させるために極めて有効であるとの知見を得て、
本発明を完成した。p e 2 + , M Oh + , Fe 3 +
We have obtained the knowledge that culturing in a medium enriched with each of these salts to contain one or more of these salts is extremely effective in increasing the amount of diacetyl produced.
The invention has been completed.
すなわち、本発明はクエン酸醗酵能を有する乳酸菌によ
りクエン酸からジアセチルを醗酵生成せしめるに際して
、前記乳酸菌のクエン酸を含有する培養基質に鉄、銅お
よびモリブデンの１種または２種以上を無機塩類または
有機塩類の形で添加することを特徴とするものである。That is, in the present invention, when producing diacetyl from citric acid by fermentation using lactic acid bacteria having citric acid fermentation ability, one or more of iron, copper, and molybdenum are added to the culture substrate containing citric acid of the lactic acid bacteria as an inorganic salt or It is characterized by being added in the form of organic salts.
本発明におけるクエン酸醗酵能を有する乳酸菌としては
、クエン酸醗酵能を有する乳酸菌であればいずれでも使
用できるが、Ｓｔｒｅｐｔｏｃｏｃｃｕｓ １ａ−ｃｔ
ｉｓ ５ｕｂｓｐ、ｄｉａｃｅｔｙｌａｃｔｉｓ及びＬ
ｅｕｃｏｎｏｓｔｏｃ ｃｒｅｍ−ｏｒｉｓがジアセチ
ル生成能が優れているので好ましい。As the lactic acid bacteria having citric acid fermentation ability in the present invention, any lactic acid bacteria having citric acid fermentation ability can be used, but Streptococcus 1a-ct
is 5ubsp, diacetylactis and L
Euconostoc crem-oris is preferred because it has an excellent ability to produce diacetyl.
培養基質として脱脂乳あるいは脱脂粉乳溶液を主に使用
する場合は、両者ともクエン酸を固形分中に約１％を含
有しているので、クエン酸を培養基質に添加しなくても
差支えないが、培養基質にクエン酸が含有していないか
、含有量の少ない場合あるいはジアセチルを多量に生成
させる必要のある場合は、所望のジアセチル生成量に応
じた量のクエン酸を培養基質に添加含有せしめる。なお
添加するクエン酸はクエン酸塩として添加するとよい。If skim milk or skim milk powder solution is mainly used as the culture substrate, since both contain about 1% citric acid in solid content, there is no problem even if citric acid is not added to the culture substrate. If the culture substrate does not contain citric acid or has a low content, or if it is necessary to generate a large amount of diacetyl, add citric acid to the culture substrate in an amount corresponding to the desired amount of diacetyl produced. . Note that the citric acid to be added is preferably added as a citrate.
本発明において培養基質に添加する鉄、銅およびモリブ
デンの無機塩類または有機塩類は単独でもよいし、組合
せて添加してもよい。その添加量は鉄イオン、銅イオン
、モリブデンイオンの合計量が０．０１ｍＭ〜１０ｍＭ
程度となるように添加することが好ましい。０．０１ｍ
Ｍより少ないとジアセチル生成量の増強の効果が認めら
れない。１０ｍＭより多いと培養基質の着色、凝固など
の現象を生ずるおそれがある。In the present invention, the inorganic or organic salts of iron, copper, and molybdenum added to the culture substrate may be added alone or in combination. The total amount of iron ions, copper ions, and molybdenum ions added is 0.01mM to 10mM.
It is preferable to add it to a certain extent. 0.01m
If it is less than M, no effect of increasing the amount of diacetyl produced will be observed. If the amount exceeds 10 mM, phenomena such as coloring and coagulation of the culture substrate may occur.
なお、醗酵孔の培養基質となる乳中には、銅。In addition, there is copper in the milk, which serves as the culture substrate for the fermentation pores.
鉄、モリブデンがそれぞれ３〜１７μｇ／Ｌｏｏｍ １
、１６〜６２μｇ／１００ｍ６ 、 ２〜１５μｇ／
１００ｍβ含まれていることが知られている（牛乳の化
学、Ｐ６４．津郷友吉ら著、地球出版）。しかしこのよ
うな培養基質の金属イオン含量では乳酸菌のクエン酸代
謝に関与する酵素系を活性化するには不十分である。Iron and molybdenum each 3-17 μg/Loom 1
, 16-62μg/100m6, 2-15μg/
It is known that it contains 100 mβ (Chemistry of Milk, p. 64, written by Tomoyoshi Tsugo et al., Chikyu Publishing). However, the metal ion content of such a culture substrate is insufficient to activate the enzyme system involved in citric acid metabolism of lactic acid bacteria.
つぎに、本発明の試験例を示す。Next, test examples of the present invention will be shown.
試験例１
Ｓｔｒｅｐｔｏｃｏｃｃｕｓ １ａｃｔｉｓ ５ｕ
ｂｓｐ、ｄｉａｃｅｔｙｌａｃｔｉｓ（八ＴＣＣ１１０
０７）のクエン酸代謝に関与する諸活性のうち、クエン
酸取込み活性、クエン酸リアーゼ活性ならびにジアセチ
ル生成活性につき検討し、第１表の結果を得た。Test Example 1 Streptococcus 1actis 5u
bsp, diacetylactis (8TCC110
Among the various activities involved in citric acid metabolism of No. 07), citric acid uptake activity, citrate lyase activity, and diacetyl production activity were investigated, and the results shown in Table 1 were obtained.
無添加 ｉｏｏ ｉｏｏ ｔｏ。Additive-free ioo ioo to.
ＣｕＣ１ｚ ・２ＨｚＯ９３２２０７４０Ｍｎ５Ｏａ、
Ｈ４Ｈｚ０ １００ １００ １３１Ｍ
ｇ５０４・７１（ｔｏ １００ １０３
１４１ＺｎＳＯｎ ・７ＨｚＯ１００１１５１３５Ｎ
ａｚＭＯＯ４・２Ｈｚ０ １０１ ９８
２９３ＦｅＳＯｎ ・７Ｈｚ０ ９１ １０
０ ２８６ＡＩＣ１，・６Ｈ，０１００１２０７３
Ｆｅｚ （Ｓｏｔ） ｓ １００ １００
２３０（１１クエン酸取込み活性測定法
５ｔｒｅρｔｏｃｏｃｃｕｓ １ａｃｔｉｓ ５ｕｂｓ
ｐ、ｄｉａｃｅｔｙｌａｃｔｉｓ（ＡＴＣＣ１１００７
）をＭＲ３培地で培養し、滅菌水にて菌体洗浄後、０．
１Ｍリン酸緩衝液（ｐＨ６，０）に懸濁する。本液９ｍ
ｆに０．２％クエン酸ナトリウムを１ｒｎｌ加え、３０
℃にて３０分間反応する。反応後０．４５μｍ（ポアサ
イズ）のフィルターにて除菌した“濾液中のクエン酸含
量を測定する。反応前後におけるクエン酸量の変化より
乾菌重量１ｇ当りの３０分間反応させた際のクエン酸の
菌体への取込み量を求める。CuC1z・2HzO93220740Mn5Oa,
H4Hz0 100 100 131M
g504・71(to 100 103
141ZnSOn ・7HzO100115135N
azMOO4・2Hz0 101 98
293FeSOn ・7Hz0 91 10
0 286AIC1,・6H,010012073
Fez (Sot) s 100 100
230 (11 Citric acid uptake activity measurement method 5 treρtococcus 1actis 5 ubs
p. diacetylactis (ATCC11007
) was cultured in MR3 medium, and after washing the bacterial cells with sterile water, 0.
Suspend in 1M phosphate buffer (pH 6,0). Main liquid 9m
Add 1rnl of 0.2% sodium citrate to f,
React for 30 minutes at ℃. After the reaction, the citric acid content in the filtrate was sterilized using a 0.45 μm (pore size) filter. From the change in the amount of citric acid before and after the reaction, the citric acid content per 1 g of dry bacteria weight after 30 minutes of reaction was determined. Determine the amount of uptake into the bacterial body.
（２）クエン酸リアーゼ活性測定法
５００ ｄのＭＲ３培地にてＳｔｒｅｐｔｏｃｏｃｃｕ
ｓ １ａｃｔｉｓｓｕｂｓｐ、ｄｉａｃｅｔｙｌａｃｔ
ｉｓ（ＡＴＣＣ１１００７）を３０℃にて２０時間培養
後、滅菌水にて２図画体洗浄を繰返し、０．１Ｍリン酸
緩衝液（ｐＨ６，０）で３０ｍＺの菌体懸濁液を調製す
る。本液を超音波処理にて菌体破砕し、２５．００Ｏｒ
、ｐ、ｔにて遠心分離し、その上澄液を一夜透析処理し
、粗酵素液とする。５ｍＭクエン酸ナトリウム、１ｍＭ
塩化マグネシウムを含むＯ，１Ｍリン酸緩衝液３−に粗
酵素液０．５−を加え、３０℃にて６０分間反応させ、
沸腋水浴中で反応を停止する。本液を３００Ｏｒ、ｐ、
ｍ、で５分間遠心分雌後、上澄液中の生成酢酸量を測定
する。クエン酸リアーゼ１単位は１μｍｏｌの酢酸を３
０℃１分間で生成させるのに必要とする酵素量とし、比
活性は酵素タンパク１■あたりの単位数とする。(2) Citrate lyase activity measurement method Streptococcus in 500 d MR3 medium
s 1 actissubsp, diacetylact
After culturing is (ATCC 11007) at 30° C. for 20 hours, the cells were washed twice with sterile water, and a 30 mZ cell suspension was prepared with 0.1 M phosphate buffer (pH 6,0). The bacterial cells of this solution were crushed by ultrasonication, and 25.00 Or
, p, and t, and the supernatant was dialyzed overnight to obtain a crude enzyme solution. 5mM sodium citrate, 1mM
Add 0.5- of the crude enzyme solution to O, 1M phosphate buffer 3- containing magnesium chloride, react at 30°C for 60 minutes,
Stop the reaction in a boiling water bath. Add this solution to 300 Or, p.
After centrifugation for 5 minutes at m, measure the amount of acetic acid produced in the supernatant. One unit of citrate lyase is equivalent to 3 units of 1 μmol of acetic acid.
This is the amount of enzyme required to produce the enzyme in 1 minute at 0°C, and the specific activity is the number of units per 1 inch of enzyme protein.
（３） ジアセチル生成活性測定法
Ｋａｎｅｋｏら（Ａｇｒｉｃ、Ｂｉｏｌ、Ｃｈｅｍ、
、５０．２６３９（１９８６））の方法に従って測定し
た。(3) Diacetyl production activity measurement method Kaneko et al. (Agric, Biol, Chem,
, 50.2639 (1986)).
第１表の結果からＣｕ”がクエン酸リアーゼ活性を、Ｃ
ｕ２２ Ｍｏ６０． Ｆｅ”、 ｐｅ３＋がジアセチル
生成活性を高めることがわかる。From the results in Table 1, Cu” increases citrate lyase activity, C
u22 Mo60. It can be seen that pe3+ increases diacetyl production activity.
試験例２
Ｃｕ”、 Ｍｏ”、 Ｆｅ”、 Ｆｅ’＋をそれぞれ１
、０ｍＭとなるように添加した１０％脱脂粉乳培地に
Ｓｔｒｅｐｔｏｃｏ−ｃｃｕｓ １ａｃｔｉｓ ５ｕｂ
ｓｐ、ｄｉａｃｅｔｙｌａｃｔｉｓ（ＡＴＣＣ１１００
７）またはＬｅｕｃｏｎｏｓｔｏｃ ｃｒｅｍｏｒｉｓ
（八ＴＣＣ１９２５４）をそれぞれ接種し、３０℃で２
０時間培養後のジアセチル生成量を測定した。結果を第
２表に示す。Test Example 2 1 each of Cu", Mo", Fe", and Fe'+
, Streptococcus 1actis 5ub in 10% skim milk powder medium added to 0mM.
sp, diacetylactis (ATCC1100
7) or Leuconostoc cremoris
(8TCC19254) and inoculated at 30℃ for 2 hours.
The amount of diacetyl produced after 0 hour of culture was measured. The results are shown in Table 2.
＊ ＊ ： Ｌｅｕｃｏｎｏｓｔｏｃ ｃｒｅｍｏｒｉ
ｓ（ＡＴＣＣ１９２５４）ジアセチル測定法
Ｋａｎｅｋｏら（八ｇｒｉｃ、Ｂｉｏ１．ｃｈｅｍ、、
５０．２６３９（１９８６））の方法に従って測定した
。＊ ＊ : Leuconostoc cremori
s (ATCC19254) diacetyl measurement method Kaneko et al. (8gric, Bio1.chem,
50.2639 (1986)).
第２表の結果から、Ｃｕ２′″１Ｍ０６″″、 Ｆｅｚ
′″ ｐ ｅ ３ ＋の添加がジアセチル生成量を高め
るために極めて有効であることがわかる。この現象は、
第１表が示すように、培地中より菌体内にとり込んだク
エン酸よりジアセチルにいたる代謝において、金属イオ
ンが有効に作用したことによるものと考えられる。From the results in Table 2, Cu2′″1M06″″, Fez
''' It can be seen that the addition of p e 3 + is extremely effective for increasing the amount of diacetyl produced. This phenomenon is
As shown in Table 1, this is thought to be due to the effective action of metal ions in the metabolism of citric acid taken into the bacterial cells from the medium to diacetyl.
実施例１
３０％脱脂粉乳溶液（０，０５％酵母エキスを含む）１
００ ｋｇに塩化第２鉄５ｇ及びクエン酸ナトリウムｌ
ｅｇを加え、９５℃にて１５分間殺菌後、３０℃に冷却
し、Ｓｔｒｅｐｔｏｃｏｃｃｕｓ １ａｃｔｔｓ ５ｕ
ｂｓｐ、ｄｉａｃｅｔｙｌａｃｔｉｓ（ＡＴＣＣ１１０
０７）を含むスターターを１．５ｋｇ接種し、これを３
０℃で１６時間醗酵させて得られた醗酵乳中のジアセチ
ル生成量は３０■／ｋｇであった。Example 1 30% skim milk powder solution (containing 0.05% yeast extract) 1
00 kg with 5 g of ferric chloride and l of sodium citrate
Streptococcus 1actts 5u was added and sterilized at 95℃ for 15 minutes, cooled to 30℃, and Streptococcus 1actts 5u
bsp, diacetylactis (ATCC110
Inoculate 1.5 kg of starter containing 07) and inoculate this with 3
The amount of diacetyl produced in the fermented milk obtained by fermentation at 0° C. for 16 hours was 30 μ/kg.
上記したように本発明によれば、銅、鉄、モリブデンを
添加するという簡単な手段により、ジアセチルを主要な
香気成分としている醗酵乳製品のジアセチル生成量を高
めることにより、より芳香のすぐれた醗酵乳製品を得る
ことができる。As described above, according to the present invention, by simply adding copper, iron, and molybdenum, the amount of diacetyl produced in fermented dairy products whose main aroma component is diacetyl is increased, resulting in fermentation with a more excellent aroma. You can get dairy products.
また、高濃度にジアセチルを生成させることにより、得
られた醗酵製品はヨーグルト、バター。In addition, by producing diacetyl in high concentrations, the fermented products obtained are yogurt and butter.
マーガリン、パンなどのフレーバー付与物質として使用
できる。It can be used as a flavoring substance in margarine, bread, etc.
Claims (4)
らジアセチルを醗酵生成せしめるに際して、前記乳酸菌
のクエン酸を含有する培養基質に鉄、銅およびモリブデ
ンの１種または２種以上を無機塩類または有機塩類の形
で添加することを特徴とする乳酸菌によるジアセチル生
成量の増強法。(1) When producing diacetyl from citric acid by fermentation using lactic acid bacteria having citric acid fermentation ability, one or more of iron, copper and molybdenum are added to the citric acid-containing culture substrate of the lactic acid bacteria as inorganic or organic salts. A method for enhancing the amount of diacetyl produced by lactic acid bacteria, characterized by adding in the form of.
ン及びモリブデンイオンの合計量が０．０１〜１０ｍＭ
となるように添加することを特徴とする特許請求の範囲
第１項記載の乳酸菌によるジアセチル生成量の増強法。(2) The total amount of iron ions, copper ions and molybdenum ions in the inorganic salts or organic salts is 0.01 to 10mM.
The method for enhancing the amount of diacetyl produced by lactic acid bacteria according to claim 1, characterized in that the amount of diacetyl produced by lactic acid bacteria is increased.
ｃｏ−ｃｕｓｌａｃｔｉｓｓｕｂｓｐ．ｄｉａｃｅｔｙ
ｌａｃｔｉｓであることを特徴とする特許請求の範囲第
１項および２項のいずれかに記載の乳酸菌によるジアセ
チル生成量の増強法。(3) Lactic acid bacteria with citric acid fermentation ability are Strepto
co-cuslactissubsp. diacety
lactis. The method for enhancing the amount of diacetyl produced by lactic acid bacteria according to any one of claims 1 and 2, characterized in that the lactic acid bacteria are L. lactis.
ｓｔｏｃｃｒｅｍｏｒｉｓであることを特徴とする特許
請求の範囲第１項及び２項のいずれかに記載の乳酸菌に
よるジアセチル生成量の増強法。(4) Lactic acid bacteria with citric acid fermentation ability are Leucono
3. The method for enhancing the amount of diacetyl produced by lactic acid bacteria according to any one of claims 1 and 2, characterized in that the lactic acid bacteria is A. stoccremoris.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP3337887A JP2568537B2 (en) | 1987-02-18 | 1987-02-18 | Method for enhancing diacetyl production by lactic acid bacteria |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP3337887A JP2568537B2 (en) | 1987-02-18 | 1987-02-18 | Method for enhancing diacetyl production by lactic acid bacteria |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPS63202390A true JPS63202390A (en) | 1988-08-22 |
| JP2568537B2 JP2568537B2 (en) | 1997-01-08 |
Family
ID=12384931
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP3337887A Expired - Lifetime JP2568537B2 (en) | 1987-02-18 | 1987-02-18 | Method for enhancing diacetyl production by lactic acid bacteria |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JP2568537B2 (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0430406A2 (en) * | 1989-11-28 | 1991-06-05 | Meiji Milk Products Company Limited | Method for the fermentative production of diacetyl and acetoin using lactic acid bacterium |
-
1987
- 1987-02-18 JP JP3337887A patent/JP2568537B2/en not_active Expired - Lifetime
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0430406A2 (en) * | 1989-11-28 | 1991-06-05 | Meiji Milk Products Company Limited | Method for the fermentative production of diacetyl and acetoin using lactic acid bacterium |
Also Published As
| Publication number | Publication date |
|---|---|
| JP2568537B2 (en) | 1997-01-08 |
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