JP5579701B2 - 乳酸菌の培養方法および飲食品 - Google Patents
乳酸菌の培養方法および飲食品 Download PDFInfo
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- JP5579701B2 JP5579701B2 JP2011507095A JP2011507095A JP5579701B2 JP 5579701 B2 JP5579701 B2 JP 5579701B2 JP 2011507095 A JP2011507095 A JP 2011507095A JP 2011507095 A JP2011507095 A JP 2011507095A JP 5579701 B2 JP5579701 B2 JP 5579701B2
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- lactic acid
- acid bacteria
- phosphate
- mass
- milk
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- BAQAVOSOZGMPRM-QBMZZYIRSA-N sucralose Chemical compound O[C@@H]1[C@@H](O)[C@@H](Cl)[C@@H](CO)O[C@@H]1O[C@@]1(CCl)[C@@H](O)[C@H](O)[C@@H](CCl)O1 BAQAVOSOZGMPRM-QBMZZYIRSA-N 0.000 description 1
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- 229910000404 tripotassium phosphate Inorganic materials 0.000 description 1
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- RYFMWSXOAZQYPI-UHFFFAOYSA-K trisodium phosphate Chemical compound [Na+].[Na+].[Na+].[O-]P([O-])([O-])=O RYFMWSXOAZQYPI-UHFFFAOYSA-K 0.000 description 1
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- HEBKCHPVOIAQTA-SCDXWVJYSA-N xylitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)CO HEBKCHPVOIAQTA-SCDXWVJYSA-N 0.000 description 1
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
- A23C9/00—Milk preparations; Milk powder or milk powder preparations
- A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
- A23C9/123—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt
- A23C9/1234—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt characterised by using a Lactobacillus sp. other than Lactobacillus Bulgaricus, including Bificlobacterium sp.
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
- A23C9/00—Milk preparations; Milk powder or milk powder preparations
- A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
- A23C9/127—Fermented milk preparations; Treatment using microorganisms or enzymes using microorganisms of the genus lactobacteriaceae and other microorganisms or enzymes, e.g. kefir, koumiss
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
- A23C9/00—Milk preparations; Milk powder or milk powder preparations
- A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
- A23C9/13—Fermented milk preparations; Treatment using microorganisms or enzymes using additives
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- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/04—Preserving or maintaining viable microorganisms
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- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/38—Chemical stimulation of growth or activity by addition of chemical compounds which are not essential growth factors; Stimulation of growth by removal of a chemical compound
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2400/00—Lactic or propionic acid bacteria
- A23V2400/11—Lactobacillus
- A23V2400/125—Casei
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- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Food Science & Technology (AREA)
- Polymers & Plastics (AREA)
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- Dairy Products (AREA)
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Description
従って、本発明は、乳酸菌数を安定して維持することができる乳酸菌培養物を得るための乳酸菌の培養方法を提供することをその目的とするものである。
そして更に、本発明は、品質安定性に優れた乳酸菌培養物を含む飲食品を得ることを目的とするものである。
また、この培養物を利用することにより、優れた品質安定性を有する発酵乳製品などの飲食品が製造できることを見出した。
また、本発明の方法は、増殖促進や生残性改善に有用な既知の物質を組み合わせることで、活性の高い乳酸菌を安定して多く含む培養物を得ることができるので、生菌数が重要視される乳酸菌飲料等の発酵乳食品の製造に特に好適である。
更に言えば、本発明の方法により、天然物である乳製品を、季節や産地、加工方法等に関係なく、乳酸菌を培養する培地の原料に供することができる。
以下、脱脂粉乳を例に挙げて、乳成分中の遊離リン酸濃度の測定方法を説明する。なお、本願明細書中においては、以下に示す方法により測定した値を乳成分中の遊離リン酸濃度と定義する。
(1)試薬
(a)アスコルビン酸溶液(72g/L)
L(+)-アスコルビン酸(和光純薬工業(株)、特級)7.2gを水に溶解して100mLとし、0〜10℃の暗所に保存する。
(b)モリブデン酸アンモニウム溶液
七モリブデン酸六アンモニウム4水和物(和光純薬工業(株)、特級)6.0gとビス[(+)タルトラト]二アンチモン(III)酸カリウム3水和物(和光純薬工業(株)、特級)0.24gを水約300mLになるように溶解し、これに硫酸(2+1)120mLを加えて500mLとする。
(c)モリブデン酸アンモニウム−アスコルビン酸混合溶液(発色液)
モリブデン酸アンモニウム溶液とアスコルビン酸溶液(72g/L)とを体積比で5:1になるように混合する(用時調製)。
(d)リン酸イオン標準原液(50μg PO4 3-‐P/mL)
リン酸二水素カリウム(pH標準液用)を105±2℃で約2時間加熱し、デシケーター中で放冷する。その0.2197gを取り、1000mLとする。0〜10℃の暗所にて保存する。
(e)リン酸イオン標準溶液(1μgPO4 3-‐P/mL)
リン酸イオン標準原液(50μgPO4 3-‐P/mL)1mLを50mL容メスフラスコで定容する。
(2)検量線
(i)水を試験管に5.0mL、4.75mL、4.5mL、4.0mL、3.0mL、0.0mLずつ入れる。
(ii)(i)の各試験管にリン酸イオン標準溶液(1μgPO4 3-‐P/mL)を0.0mL、0.25mL、0.5mL、1.0mL、2.0mL、5.0mLそれぞれ加え、全量5.0mLとする。
(iii)発色液を400μL加えて、約15分間放置する。
(iv)30分以内に、分光光度計にてUV880nmの吸光度を測定する。
(3)測定手順
(i)試料(脱脂粉乳2.0g)を量り、水(または温水)で溶解後、100mL容メスフラスコで定容し、1時間以上放置する。
(ii)(i)の約6mLをビバスピン6(5,000MWCO)に量り取り、遠心分離機で限外ろ過する(7,500G、30分、25℃)。
(iii)(ii)の通過液を1mLホールピペットで正確に100mL容メスフラスコに量り取り、水で定容する。
(iv)(iii)の溶液を5mLホールピペットで試験管に量り取る。
(v)発色液を400μL加えて、約15分間放置する。
(vi)30分以内に分光光度計にてUV880nmの吸光度を測定する。
(vii)(2)で求めた検量線から遊離リン酸量(μg/Vial)を求める。求めた遊離リン酸量を用いて下記の式から脱脂粉乳中に含まれる遊離リン酸の割合を求める。
脱脂粉乳中の遊離リン酸%
=遊離リン酸量(μg/Vial)×10,000mL/5mL×100g/2g×1g/1,000,000μg
※分析方法は、食品成分試験法およびJIS K0102(工場排水試験法、1998)のリン酸イオンの分析法(モリブデン青(アスコルビン酸還元)吸光光度法)を参考に行った。
ここで、無脂乳固形分(SNF)あたりのタンパク質含有量は、下記式1により算出することができる。
一方で、本発明において、前記した乳成分と共に乳酸菌を培養する培地に用いるリン酸塩としては、水溶性のリン酸塩、具体的には、リン酸二水素ナトリウム、リン酸水素二ナトリウム、リン酸二水素アンモニウム、リン酸水素二アンモニウム、リン酸二水素カリウム、リン酸水素二カリウム、リン酸三ナトリウム、リン酸三カリウム等を好ましいものとして挙げることができる。これらのリン酸塩は、2種以上を組み合わせて、培地のpHが中性領域(pH6〜8)となるように添加することが好ましい。
なお、培地における乳成分の濃度は、特に制限されるわけではなく、一般的な濃度とすればよく、例えば、5質量%〜30質量%、好ましくは、10質量%〜20質量%程度である。
すなわち、本発明において、リン酸塩の使用量は、下記式2により算出して設定することができる。なお、このとき、リン酸塩の使用量は、リン酸塩中のリン酸が全て遊離リン酸とした場合の濃度として設定することができる。
(1)遊離リン酸濃度が0.20質量%の乳成分を用いる場合
・下限値
(0.25−0.20)×20/100×155/31=0.05%
・上限値
(0.50−0.20)×20/100×155/31=0.30%
すなわち、遊離リン酸濃度が0.20質量%の乳成分を用いて、20質量%の培地を調製する場合には、リン酸塩を0.05質量%以上、好ましくは0.05質量%〜0.30質量%の範囲で添加すればよい。つまり、乳成分を20質量%含む培地100gに関して、上記リン酸塩を0.05g以上、好ましくは0.05g〜0.30g添加すればよい。
(2)遊離リン酸濃度が0.24質量%の乳成分を用いる場合
・下限値
(0.25−0.24)×20/100×155/31=0.01%
・上限値
(0.50−0.24)×20/100×155/31=0.26%
すなわち、遊離リン酸濃度が0.24質量%の乳成分を用いて、20質量%の培地を調製する場合には、リン酸塩0.01質量%以上、好ましくは0.01質量%〜0.26質量%の範囲で添加すればよい。つまり、乳成分を20質量%含む培地100gに関して、上記リン酸塩を0.01g以上、好ましくは0.01g〜0.26g添加すればよい。
一方で、遊離リン酸濃度が0.50質量%である乳成分を基本原料として培地を調製した場合における当該培地中の理論上の遊離リン酸濃度よりも高くなると、得られる乳酸菌発酵物における乳酸菌数の安定化は向上するが、これを含む飲食品において、保存による凝集や沈殿など、品質の劣化を生じることがある。
(乳成分の成分分析)
オーストラリアを原産国とするMurray Goulburn社製の脱脂粉乳(以下、単に「試料」という。)について、以下により遊離リン酸濃度とタンパク質含有量を測定した。
本願明細書中、段落[0015]に記載の方法に従って測定を行った。
その結果、遊離リン酸濃度は、0.23%であった。
以下の分析値を用いて本願明細書中、段落[0017]に記載の式1により算出した。
脱脂粉乳中のタンパク質含有量:32.8%
水分含有量:3.8%、脂質含有量:0.6%
その結果、無脂乳固形分当たりのタンパク質含有量は、34.3%/SNFであった。
(乳酸菌培養物の調製)
試験例1の試料を用いて、表1に示すとおりの組成で脱脂粉乳、リン酸塩、ぶどう糖を水に溶解し培地を調製した。なお、リン酸塩の使用量は、リン酸塩中のリン酸が全て遊離リン酸とした場合の濃度とした。(以下、同様とする。)この培地を100℃で90分間殺菌し、次いで、ラクトバチルス・カゼイを0.5%接種し、pH3.6程度になるまで培養して得た乳酸菌培養物の培養終了時のpHと乳酸菌数を測定した。また、この培養物600mLにぶどう糖果糖液糖400mL及び滅菌水1.5Lを加えて均質化し、乳酸菌飲料(実施品、比較品)を製造した。製造直後と10℃で14日間保存後のpH、乳酸菌数を測定した。その結果を下表1に示す。
(乳酸菌飲料の製造)
試験例1の試料を用いて、表2に示すとおりの組成で脱脂粉乳、リン酸塩、ぶどう糖を水に溶解し培地を調製した。これらの培地を100℃で90分間殺菌し、ラクトバチルス・カゼイを0.5%接種し、pH3.6程度になるまで培養して得た乳酸菌培養物(A〜C)の培養終了時のpHと乳酸菌数を測定した。その結果を表2に示す。次に、この培養物600mLにぶどう糖果糖液糖400mL及び滅菌水1.5Lを加えて均質化し、65mL容容器に充填して乳酸菌飲料(実施品1〜3)を製造した。製造直後と10℃で14日間保存後のpH、乳酸菌数を測定した。また、保存後の乳酸菌飲料については、沈殿量を目視により確認するとともに、沈殿量とホエーオフ(離水量)を測定した。その結果を表3に示す。
なお、目視による沈殿量の評価は、下記の指標に基づいて判定した。
1.沈殿
++ : 非常に有り
+ : 有り
± : 僅かに有り(製品として問題無い程度)
− : 無し
D−C/A−C
D;容器+沈殿物の重さ
(製品保存後の内容物を静かに捨て、1分間倒置後の沈殿物が付着した容器の重さ)
C;容器の重さ
A;容器+充填液の重さ
また、得られた乳酸菌飲料は、風味も良好で、保存後においても沈殿や分離なども生じることはなく、品質安定性にも優れていた。
Claims (7)
- 前記乳成分の無脂乳固形分(SNF)あたりのタンパク質含有量が35質量%未満である、請求項1に記載の乳酸菌の培養方法。
- 前記リン酸塩が水溶性である、請求項1又は2に記載の乳酸菌の培養方法。
- 前記乳成分が脱脂粉乳である、請求項1乃至3のいずれか一項に記載の乳酸菌の培養方法。
- 前記乳酸菌がラクトバチルス・カゼイである、請求項1乃至4のいずれか一項に記載の乳酸菌の培養方法。
- 請求項1乃至5のいずれか一項に記載の乳酸菌の培養方法を含む、発酵乳製品の製造方法。
- 請求項6に記載の発酵乳製品を原料として用いることを含む、乳酸菌飲料の製造方法。
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