AU2011327288B2 - Lactobacillus helveticus having high proteolysis activity - Google Patents
Lactobacillus helveticus having high proteolysis activity Download PDFInfo
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- AU2011327288B2 AU2011327288B2 AU2011327288A AU2011327288A AU2011327288B2 AU 2011327288 B2 AU2011327288 B2 AU 2011327288B2 AU 2011327288 A AU2011327288 A AU 2011327288A AU 2011327288 A AU2011327288 A AU 2011327288A AU 2011327288 B2 AU2011327288 B2 AU 2011327288B2
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- milk
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- lactic acid
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Classifications
-
- 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
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/135—Bacteria or derivatives thereof, e.g. probiotics
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/17—Amino acids, peptides or proteins
- A23L33/18—Peptides; Protein hydrolysates
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- 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
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- 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
- C12N1/205—Bacterial isolates
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P13/00—Preparation of nitrogen-containing organic compounds
- C12P13/04—Alpha- or beta- amino acids
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P21/00—Preparation of peptides or proteins
- C12P21/06—Preparation of peptides or proteins produced by the hydrolysis of a peptide bond, e.g. hydrolysate products
-
- 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
- A23V2400/147—Helveticus
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/225—Lactobacillus
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Genetics & Genomics (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Food Science & Technology (AREA)
- Polymers & Plastics (AREA)
- Medicinal Chemistry (AREA)
- Virology (AREA)
- Nutrition Science (AREA)
- Molecular Biology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Tropical Medicine & Parasitology (AREA)
- Mycology (AREA)
- Biomedical Technology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Dairy Products (AREA)
- Coloring Foods And Improving Nutritive Qualities (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Peptides Or Proteins (AREA)
Abstract
The present invention provides a lactic acid bacterium which has high proteolysis activity and thus has excellent peptide production ability and/or amino acid production ability. The present invention also provides fermented milk which contains an amino acid and/or a peptide at high concentration and has a good flavor. The present invention specifically provides a lactic acid bacterium belonging to the genus
Description
DESCRIPTION LACTOBACILLUS HELVETICUS HAVING HIGH PROTEOLYSIS ACTIVITY TECHNICAL FIELD [0001] The present invention relates to a novel lactic acid bacterium belonging to Lactobacillus helveticus. The present invention further relates to fermented milk obtained by fermenting milk with the lactic acid bacterium, and a composition or a food or drink product comprising the lactic acid bacterium. The present invention further relates to a method for producing amino acids and/or peptides using the lactic acid bacterium. BACKGROUND ART [0002] Milk proteins and food proteins are known to contain functional peptides latently. The use of milk proteins or food proteins that have been processed by enzymatic degradation or fermentation has been attempted. For example, as such functional peptides, the tripeptides VPP and IPP have been reported to have an angiotensin converting enzyme (ACE) inhibiting activity and thus exhibit a hypotensive effect (Patent Literatures 1-4 and Non-patent Literature 1). [0003] As a milk-derived functional peptide other than VPP and IPP, YP, which is a dipeptide, has been reported to have hypotensive and anxiolytic effects (Patent Literature 5). It has also been reported that the dipeptides IP, AP, EP, RP, QP, TP, MP, and G? are of a less degradable peptide group having ACE inhibiting activity (e.g., Patent Literature 6). [0004] Meanwhile, Lactobacillus helveticus (L. helveticus) has been conventionally used for producing dairy products and is known to have high proteolysis activity. To provide fermented milk that is more easily ingested and processed, some L. helveticus strains that have high proteolysis activity and good peptide productivity have been reported (Patent Literatures 1-4 and 7, Non-patent Literature 1). [0005] Proteins are composed of amino acids, and thus can be the source of amino acids supplied through promotion of proteolysis. Amino acids are known to have various functions in addition to nutrition and flavor. For example, since branched chain amino acids (BCAAs) are contained in proteins composing muscle, in which BCAAs are consumed upon exercise or - 1 the like and used as energy, it is known that supplying the BCAAs is important. Aalanine (Ala) is a glycogenic amino acid known to serve as an energy source for the liver. Since Ala consumes NADH that is produced upon alcohol degradation, it is suggested that the degradation reaction in the liver proceeds in the presence of Ala. It is further known that Ala induces the taste sensation of umami (i.e., good taste and flavor) and has shionare and sunare effects (i.e., effects of lowering the sensations of salty and sour tastes) and an effect of masking acrid taste. Lysine (Lys) is said to be the amino acid that is the most easily deficient among the essential amino acids. Lys has to be ingested as a raw material of antibodies, hormones, enzymes, or the like. It has been reported that Lys is also involved in calcium absorption, collagen fonnation, and bone tissue production. Serine (Ser) is important as a precursor for various important substances in the body, such as phosphatidyl serine and other amino acids. It is also known that Ser suppresses the decrease in blood cholesterol level or the brain cell death through neuron activation in the braine, thereby preventing brain aging and serving as a skin moisturizing ingredient. [0006] For production of such amino acids, microoragnisms have been aggressively used. Nevertheless, there are not many examples of amino acids that can be ingested directly in the form of products fermented by microorganisms. Regarding lactic acid bacteria, lactic acid bacteria having good activity to degrade to amino acids have been screened for in order to improve the flavor of Japanese pickles. However, there are no known examples of examining the direct use of fermented milk in order to ingest functional amino acids and functional peptides. Prior Art Literatures Patent Literature [0007] Patent Literature 1: European Patent No. 101 6709B Patent Literature 2: International Patent Publication WO 01/32836 Patent Literature 3: International Patent Publication WO 2007/0965 10 Patent Literature 4: European Patent No. 1820850 Patent Literature 5: European Patent No. 0821968 Patent Literature 6: European Patent No. 1661909 Patent Literature 7: JP Patent Publication (Kokai) No. 07-274949 A (1995) Non-patent Literature [0008] Non-patent Literature 1: Nakamura, Y. et al., J. Dairy Sci. Vol. 78, pp. 1253-1257, 1995 -2- SUMMARY OF THE INVENTION PROBLEM TO BE SOLVED BY THE INVENTION [0009] Therefore, an object of the present invention is to provide a lactic acid bacterium which has high proteolysis activity and good peptide production ability and/or amino acid production ability. Another object of the present invention is to provide fermented milk containing amino acids and/or peptides at high concentrations and having good flavor. MEANS FOR SOLVING THE PROBLEM [0010] As a result of intensive studies to achieve the above objects, the present inventors have succeeded in obtaining a bacterial strain having good amino acid production ability and good functional peptide production ability from milk proteins and having proteolysis activity higher than that of conventional lactic acid bacteria, from among strains of lactic acid bacteria belonging to Lactobacillus helveticus. Furthermore, the present inventors have obtained findings that fermented milk and a composition in which the amino acid and/or peptide content is high is obtained with the use of the bacterial strain, thus allowing amino acids and/or peptides to be produced. As a result, the present inventors have completed the present invention. [0011] Specifically, the present invention encompasses the following [1] to [32]. [1] A lactic acid bacterium belonging to Lactobacillus helveticus, having at least one of the following properties (a) to (d) of: (a) producing 25 imol/ml or more of free amino acids by fermenting animal milk; (b) producing 8.5 mg/ml or more of peptides by fermenting animal milk; (c) producing XP peptides and/or XPP peptids in a total amount of 260 pg/ml or more by fermenting animal milk; and (d) producing 6.0 mg/g or more of IPP and VPP peptides, which amount is converted to the amount of VPP, per acidity of fermented milk that is obtained by fermenting animal milk at 30-34*C. [2] The lactic acid bacterium according to [1], wherein 15% or more of the free amino acids are branched chain amino acids. [3] The lactic acid bacterium according to [1] or [2], wherein the free amino acids comprise at least one amino acid selected from alanine, lysine, and serine. [4] The lactic acid bacterium according to any one of [1] to [3], which produces YP peptide in an amount of 60 pg/ml or more. -3 - [5] The lactic acid bacterium according to any one of [1] to [4], which is of a mutant strain of Lactobacillus helveticus CM4 strain (Accession No. FERM BP-6060). [6] The lactic acid bacterium according to any one of [1] to [5], which is Lactobacillus helveticus CP3232 strain (Accession No. FERM BP-1 1271) or a mutant strain thereof. [7] Lactobacillus helveticus CP3232 strain (Accession No. FERM BP-1 1271). [0012] [8] A composition for producing amino acids and/or peptides from animal milk, comprising at least one lactic acid bacterium belonging to Lactobacillus helveticus. [9] The composition according to [8], which produces 25 jimol/ml or more of free amino acids by fermenting animal milk. [10] The composition according to [9], wherein 15% or more of the free amino acids are branched chain amino acids. [11] The composition according to [9] or [10], wherein the free amino acids compeises at least one amino acid selected from alanine, lysine, and serine. [12] The composition according to any one of [8] to [11], which produces peptides in an amount of 8.5 mg/ml or more by fermenting animal milk. [13] The composition according to any one of [8] to [12], which produces XP peptides and/or XPP peptides in a total amount of 260 ptg/ml or more by fermenting animal milk. [14] The composition according to any one of [8] to [13], which produces YP peptide in an amount of 60 pig/ml or more by fermenting animal milk. [15] The composition according to any one of [8] to [14], which produces 6.0 mg/g or more of IPP and VPP peptids, which amount is converted to the amount of VPP, per acidity of fermented milk that is obtained by fermenting animal milk at 30-34*C. [16] The composition according to any one of [8] to [15], wherein the lactic acid bacterium is of any one of [1] to [7]. [0013] [17] Fermented milk, which is obtained by fermenting animal milk with at least one lactic acid bacterium belonging to Lactobacillus helveticus and contains at least one of the following ingredients (a) to (d): (a) 25 ptmol/ml or more of free amino acids; (b) 8.5 mg/ml or more of peptides; (c) 260 pg/ml or more in total of XP peptides and/or XPP peptides; and (d) 6.0 mg/g or more of IPP and VPP peptides per acidity when converted to the amount of VPP. [18] The fermented milk according to [17], containing at least one lactic acid bacterium -4belonging to Lactobacillus helveticus. [19] The fermented milk according to [17] or [18], comprising at least one lactic acid bacterium of any one of [1] to [7]. [20] The fermented milk according to any one of [17] to [19], wherein 15% or more of the free amino acids are branched chain amino acids . [21] The fermented milk according to any one of [17] to [20], wherein the free amino acids comprise at least one amino acid selected from alanine, lysine, and serine. [22] The fermented milk according to any one of [17] to [21], comprising 60 jig/ml or more of YP peptide. [23] The fermented milk according to any one of [17] to [22], wherein the fermentation is performed at a temperature of 30-34*C. [24] The fermented milk according to any one of [17] to [23], for use in a hypotensive drug, a brain function improving agent, an autonomic nerve regulating agent, a parasympathetic nerve regulating agent, a sympathetic nerve regulating agent, a nutritional supplement, an energy supplement, an agent for suppressing muscle degradation and muscle damage, a muscle booster, an agent for alleviating fatigue, an endurance improving agent, a flavor improving agent, or an alcohol metabolizing agent. [25] A food or drink product, comprising at least one lactic acid bacterium of any one of [1] to [7] and/or a treated product thereof, and/or the fermented milk of any one of [17] to [24] and/or a treated product thereof. [0014] [26] A method for producing an amino acid(s) and/or a peptide(s), comprising fermenting animal milk or milk proteins using at least one lactic acid bacterium of any one of [1] to [7] or the composition of any one of [8] to [16], and then collecting the amino acid(s) and/or the peptide(s) from the obtained fermented product. [27] The method according to [26], wherein the amino acid is a branched chain amino acid. [28] The method according to [26] or [27], wherein the amino acids comprise at least one amino acid selected from alanine, lysine, and serine. [29] The method according to any one of [26] to [28], wherein the peptide is at least one peptide selected from the group consisting of XP and XPP peptides. [30] The method according to any one of [26] to [29], wherein the fermentation is performed at a temperature of 30-34*C. [0015] [31] A method for obtaining a lactic acid bacterium, which has proteolysis activity and belongs to Lactobacillus helveticus, comprising the following steps of: -5 - (a) preparing mutant strains of Lactobacillus helveticus CM4 strain (Accession No. FERM BP-6060) or Lactobacillus helveticus CP3232 strain (Accession No. FERM BP-1 1271); and (b) selecting from the mutant strains, a strain having at least one of the following properties (a) to (d) of: (a) producing 25 tmol/ml or more of free amino acids by fermenting animal milk; (b) producing 8.5 mg/ml or more of peptides by fermenting animal milk; (c) producing 260 gg/ml or more in total of XP peptide and/or XPP peptide by fermenting animal milk; and (d) producing 6.0 mg/g or more of IPP and VPP peptides, which amount is converted to the amount of VPP, per acidity of fermented milk that is obtained by fermenting animal milk at 30-34 0 C. [0016] [32] A method for producing a functional food or drink product, comprising the steps of preparing at least one lactic acid bacterium of any one of [1] to [7] and/or a treated product thereof, and/or the fermented milk of any one of [17] to [24] and/or a treated product thereof, and blending the lactic acid bacterium and/or the treated product thereof, and/or the fermented milk and/or the treated product thereof, into a food or drink product. EFFECT OF THE INVENTION [0017] According to the present invention, lactic acid bacteria belonging to Lactobacillus helveticus, having high proteolysis activity and good peptide production ability and amino acid production ability are provided. Amino acids and/or peptides produced by the lactic acid bacteria have various functions. For example, peptides such as IPP, VPP, and YP have a hypotensive effect. Furthermore, through the use of the lactic acid bacteria for fermentation, fermented milk containing a wide variety of amino acids and/or peptides having various functions at high concentrations can be obtained. The amount of IPP and VPP peptides per unit of acidity of the thus obtained fermented milk is high, so that there is no problem caused by high acidity that results from lactic acid produced as conventional fermentation proceeds. Specifically, fermented milk containing amino acids and/or peptides at levels higher than that of lactic acid produced can be produced. Therefore, with the use of a small amount of fermented milk, a product in a form having good flavor and enabling easy drinking or eating can be conveniently produced while keeping its functionality. Furthermore, the product can be provided to consumers. -6- [00 17A] According to the invention there is also provided an isolated lactic acid bacterium belonging to Lactobacillus helveticus, having the following properties: (a) producing 25 tmol/ml or more of free amino acids by fermenting animal milk; and (b) producing 8.5 mg/ml or more of peptides by fermenting animal milk; and (c) producing XP peptides and XPP peptides in a total amount of 260 tg/ml or more by fermenting animal milk; and/or (d) producing 6.0 mg/g or more of IPP and VPP peptides, which amount is converted to the amount of VPP, per acidity of fermented milk that is obtained by fermenting animal milk at 30-34 0 C. [00 17B] According to the invention there is also provided Lactobacillus helveticus CP3232 strain (Accession No. FERM BP- 11271). [00 17B] According to the invention there is also provided a composition for producing amino acids and/or peptides from animal milk, comprising at least one lactic acid bacterium belonging to Lactobacillus helveticus having the following properties: (a) produces 25 tmol/ml or more of free amino acids by fermenting animal milk; and (b) produces peptides in an amount of 8.5 mg/ml or more by fermenting animal milk; and (c) produces XP peptides and XPP peptides in a total amount of 260 ptg/ml or more by fermenting animal milk; and/or (d) produces YP peptide in an amount of 60 Lg/ml or more by fermenting animal milk; and/or (e) produces 6.0 mg/g or more of IPP and VPP peptides, which amount is converted to the amount of VPP, per acidity of fermented milk that is obtained by fermenting animal milk at 30-34 0 C. [0017C] According to the invention there is also provided fermented milk, which is obtained by fermenting animal milk with at least one lactic acid bacterium belonging to Lactobacillus helveticus having the following properties: (a) 25 [tmol/ml or more of free amino acids; and (b) 8.5 mg/mi or more of peptides; and (c) 260 pig/ml or more in total of XP peptides and XPP peptides; and/or (d) 6.0 mg/g or more of IPP and VPP peptides per acidity when converted to the amount of VPP. - 6A - [00 17D] According to the invention there is also provided a culture comprising at least one lactic acid bacterium belonging to Lactobacillus helveticus having the following properties: (a) produces 25 pLmol/ml or more of free amino acids by fermenting animal milk; and (b) produces peptides in an amount of 8.5 mg/ml or more by fermenting animal milk; and (c) produces XP peptides and XPP peptides in a total amount of 260 [g/ml or more by fermenting animal milk; and/or (d) produces YP peptide in an amount of 60 ig/ml or more by fermenting animal milk; and/or (e) produces 6.0 mg/g or more of IPP and VPP peptides, which amount is converted to the amount of VPP, per acidity of fermented milk that is obtained by fermenting animal milk at 30-34 0 C. [00 17E] According to the invention there is also provided an isolated lactic acid bacterium belonging to Lactobacillus helveticus, having the following properties: (a) producing 25 tmol/ml or more of free amino acids by fermenting animal milk; and (b) producing 8.5 mg/ml or more of peptides by fermenting animal milk; and (c) at least one of: (i) producing XP peptides and XPP peptides in a total amount of 260 tg/ml or more by fermenting animal milk; and (ii) producing 6.0 mg/g or more of IPP and VPP peptides, which amount is converted to the amount of VPP, per acidity of fermented milk that is obtained by fermenting animal milk at 30-34'C. [0017F] According to the invention there is also provided a composition for producing amino acids and/or peptides from animal milk, comprising at least one lactic acid bacterium belonging to Lactobacillus helveticus having the following properties: (a) produces 25 Itmol/ml or more of free amino acids by fermenting animal milk; and (b) produces peptides in an amount of 8.5 mg/ml or more by fermenting animal milk; and (c) at least one of: (i) produces XP peptides and XPP peptides in a total amount of 260 [tg/ml or more by fermenting animal milk; - 6B - (ii) produces YP peptide in an amount of 60 tg/ml or more by fermenting animal milk; and (iii) produces 6.0 mg/g or more of IPP and VPP peptides, which amount is converted to the amount of VPP, per acidity of fermented milk that is obtained by fermenting animal milk at 30-34'C. [0017G] According to the invention there is also provided a culture comprising at least one lactic acid bacterium belonging to Lactobacillus helveticus having the following properties: (a) produces 25 pimol/ml or more of free amino acids by fermenting animal milk; and (b) produces peptides in an amount of 8.5 mg/ml or more by fermenting animal milk; and (c) at least one of: (i) produces XP peptides and XPP peptides in a total amount of 260 tg/ml or more by fermenting animal milk; (ii) produces YP peptide in an amount of 60 .tg/ml or more by fermenting animal milk; and (iii) produces 6.0 mg/g or more of IPP and VPP peptides, which amount is converted to the amount of VPP, per acidity of fermented milk that is obtained by fermenting animal milk at 30-34*C. [0017H] According to the invention there is also provided fermented milk, which is obtained by fermenting animal milk with at least one lactic acid bacterium belonging to Lactobacillus helveticus having the following properties: (a) 25 pmol/ml or more of free amino acids; and (b) 8.5 mg/ml or more of peptides; and (c) at least one of: (i) 260 g/ml or more in total of XP peptides and XPP peptides; and (ii) 6.0 mg/g or more of IPP and VPP peptides per acidity when converted to the amount of VPP. [00171] Throughout this specification the word "comprise", or variations such as "comprises" or "comprising", will be understood to imply the inclusion of a stated element, integer or step, or group of elements, integers or steps, but not the exclusion of any other element, integer or step, or group of elements, integers or steps. -6C - [00 17J] Any discussion of documents, acts, materials, devices, articles or the like which has been included in the present specification is not to be taken as an admission that any or all of these matters form part of the prior art base or were common general knowledge in the field relevant to the present disclosure as it existed before the priority date of each claim of this application. BRIEF DESCRIPTION OF THE DRAWINGS [0018] - 6D - Fig. I shows graphs showing the amount of each amino acid (nmol/ml) contained in the supernatant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 32"C (A) or 37*C (B) for 24 hours. Fig. 2 shows graphs showing the total amount of free amino acids (gmol/ml) (A) and the amount of branched chain amino acids (gmol/ml) (B) contained in the supernatant of fermented milk obtained by fermenting milk with each of the lactic acid bacterial strains at 32*C or 37 0 C for 24 hours. Fig. 3 is a graph showing the amount of peptides (mg/ml) in the supernatant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 32*C or 37"C for 24 hours. Fig. 4 shows graphs showing the amounts of functional peptides (gg/ml) contained in the supernatant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 32*C (A) or 37 0 C (B) for 24 hours. Fig. 5 shows graphs showing the amounts of XP peptides and XPP peptides (jig/ml) contained in the supernatant of fermented milk obtained by fennenting milk with each of the indicated lactic acid bacterial strains at 32*C (A) or 37 0 C (B) for 24 hours. Fig. 6 shows graphs showing the amount of each amino acid (mnol/ml) contained in the supernatant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 32*C (A) or 37*C (B) for 24 hours. Fig. 7 shows graphs showing the total amount of free amino acids (pLmol/ml) (A) and the amount of branched chain amino acids ([tmol/ml) (B) contained in the supernatant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 32 0 C or 37*C for 24 hours. Fig. 8 is a graph showing the amount of peptides (mg/ml) in the supernatants of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 32 0 C or 37*C for 24 hours. Fig. 9 shows graphs showing the amounts of functional peptides (pg/ml) contained in the supernatant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 32*C (A) or 37*C (B) for 24 hours. Fig. 10 shows graphs showing the amounts of XP peptides and XPP peptides (jg/ml) contained in the supernatant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 32*C (A) or 37*C (B) for 24 hours. Fig. 11 shows the electrophoretic pattern of fragments obtained through amplification of genomic DNA from each of the indicated lactic acid bacterial strains by the polymerase chain reaction (PCR) method. -7- Fig. 12 shows the electrophoretic pattern of fragments obtained through amplification of genomic DNA from each of the indicated lactic acid bacterial strains by the polymerase chain reaction (PCR) method. Embodiment for Carrying Out the Invention [0019] The present invention is hereafter described in greater detail. This application claims the priority from Japanese patent application No. 2010-250754 filed on November 9, 2010, and encompasses the contents described in the description and/or drawings of this application. [0020] The lactic acid bacterium belonging to Lactobacillus helveticus according to the present invention is a lactic acid bacterium producing lactic acid from sugars via fermentation, and is also referred to as "the lactic acid bacterium (bacteria) of the present invention" herein. The lactic acid bacterium of the present invention is characterized by having high proteolysis activity and good amino acid production ability and/or good functional peptide production ability. Specifically, the lactic acid bacterium of the present invention has at least one of the following properties (a) to (d) of: (a) producing 25 pmol/ml or more, preferably 30 kmol/ml or more, of free amino acids by fermenting animal milk; (b) producing 8.5 mg/ml or more, preferably 10 mg/ml or more, of peptides by fermenting animal milk; (c) producing XP peptides and/or XPP peptides in a total amount of 260 pLg/ml or more, preferably 280 pg/ml or more, further preferably 300 [g/ml or more, by fermenting animal milk; and (d) producing 6.0 mg/g or more of IPP and VPP peptides, which amount is converted to the amount of VPP, per acidity of fermented milk obtained by fermenting animal milk at 30-34 0 C. [0021] The above properties (a) to (d) can be determined by a method known in the art, as described later in Examples, for example. Briefly, the properties (a) to (d) can be determined by fermenting milk proteins (e.g., animal milk) as substrate with a lactic acid bacterium belonging to Lactobacillus helveticus at 30-45*C, preferably at 30-34*C, for 10-30 hours and then measuring the amount of free amino acids or peptides contained in the obtained fermented product from the milk proteins, the acidity, and the like. [0022] Regarding (a), the amount of amino acids can be measured using a known amino acid -8analyzer, such as high performance liquid chromatography apparatus. The lactic acid bacterium of the present invention can produce 25 pimol/ml or more, preferably 30 jimol/ml or more, of free amino acids by fermenting animal milk. Additionally, branched chain amino acids is produced in an amount of 5 pimol/ml or more and more preferably 6 or 7 ptmol/ml or more, and preferably the branched chain amino acids (valine, leucine, and isoleucine) are 15% or more, more preferably 19% or 20% or more, of the produced free amino acids. Free amino acids preferably contain 2000 pg/ml or more of at least one amino acid selected from alanine, lysine, and serine, and more preferably contain 1500 pLg/ml each of at least two amino acids. [0023] Regarding (b), the amount of peptides can be measured by a method known in the art, such as the OPA method (Church et al., J. Dairy Sci. 1983. 66: 1219-1227). The lactic acid bacterium of the present invention can produce 8.5 mg/ml or more, preferably 10 mg/ml or more, of peptides by fermenting animal milk. [0024] Regarding (c), the term "XP peptide" refers to a peptide composed of Xaa-Pro (where Xaa denotes an arbitrary amino acid). Examples of the XP peptide include YP (Tyr-Pro), VP (Val-Pro), IP (Ile-Pro), AP (Ala-Pro), EP (Glu-Pro), RP (Arg-Pro), QP (Gln-Pro), TP (Thr-Pro), MP (Met-Pro), and GP (Gly-Pro). Furthermore, the term "XPP peptide" refers to a peptide composed of Xaa-Pro-Pro (where Xaa denotes an arbitrary amino acid). Examples of the XPP peptide include VPP (Val-Pro-Pro), IPP (Ile-Pro-Pro), and LPP (Leu-Pro-Pro). In addition, as used herein, the XP peptide and the XPP peptide- are referred to as a "dipeptide" and a "tripeptide" respectively, or are taken together to be referred to as "functional peptides." The amount of the XP peptide and/or the XPP peptide can be measured using a known LC/MS apparatus. The lactic acid bacterium of the present invention produces the XP peptide and/or the XPP peptide in a total amount of 260 pLg/ml or more, preferably 280 jig/ml or more, and more preferably 300 pg/ml or more by fermenting animal milk. In particular, 60 sig/ml or more, more preferably 65 pg/ml or more, and further preferably 70 pg/ml or more of the YP peptide is preferably produced. [0025] Regarding (d), the acidity (i.e., the concentration of acid in fermented milk) can be measured by a known neutralization titration method using a known automatic titrator or the like. Furthermore, an IPP peptide and a VPP peptide can be measured by the method described concerning (c). The lactic acid bacterium of the present invention produces 5.5 mg/ml or more, preferably 6.0 mg/ml or more, of the IPP and VPP peptides, which amount is -9converted to the amount of VPP, per acidity of fermented milk obtained by fermenting animal milk at 30-34*C. In addition, this amount is calculated using 1.02 as the specific gravity of fermented milk. Furthermore, the amount of the IPP and VPP peptides (as described herein) is calculated by the following formula in terms of the amount of VPP, which is based on ACE-inhibiting activity: (amount calculated in terms of the amount of VPP (jig/ml)) = amount of IPP (jig/ml) x 1.7 + amount of VPP (pig/ml) [0026] The lactic acid bacterium of the present invention has at least one of the properties (a) to (d) above and preferably exhibits 2, 3, or all properties above. For example, the lactic acid bacterium of the present invention has property (a), (b), (c), or (d), or have properties (a) and (b), (a) and (c), (a) and (d), (b) and (c), (b) and (d), or (c) and (d), properties (a), (b), and (c), (a), (b), and (d), (a), (c), and (d), (b), (c), and (d), or properties (a) to (d). [0027] In the present invention, any lactic acid bacteria belonging to Lactobacillus helveticus exhibiting at least one of the above properties are included in the lactic acid bacterium of the present invention. A preferable example of Lactobacillus helveticus having such properties is the Lactobacillus helveticus CP3232 strain which was screened by the present inventors themselves from mutant strains obtained using the Lactobacillus helveticus CM4 strain (Accession No. FERM BP-6060, Patent Literature 1) as a parent strain. In addition, the Lactobacillus helveticus CM4 strain and CP3232 strain were deposited by the present applicant under FERM BP-6060 and FERM BP-l 1271 as of August 15, 1997, and August 4, 2010, respectively, with the International Patent Organism Depositary, the National Institute of Advanced Industrial Science and Technology (AIST) (Tsukuba Central 6, 1-1-1 Higashi, Tsukuba, Ibaraki, Japan, 305-8566), which is an international depository authority (IDA) established under the Budapest Treaty for deposition of patent microorganisms. The Lactobacillus helveticus CP3232 strain was confirmed to have all the properties (a) to (d) above, as described in the Examples. [0028] Furthermore, the Lactobacillus helveticus CP3232 strain is a mutant strain derived from the Lactobacillus helveticus CM4 strain as a parent strain described above. Accordingly, mutant strains of the Lactobacillus helveticus CM4 strain and the Lactobacillus helveticus CP3232 strain are likely to have any of the properties (a) to (d) above. Such a mutant strain from the Lactobacillus helveticus CP3232 strain or the Lactobacillus helveticus CM4 strain is also encompassed in the present invention, as long as it has the above properties. - 10 - [0029] In the present invention, the term "mutant strain" refers to an arbitrary strain obtained from a parent strain. Specifically, it refers to a strain that is obtained from a parent strain by a method for increasing the frequency of natural mutation or artificial mutation via mutagenesis with chemical or physical mutagen, or a specific mutagenesis technique (e.g., genetic recombination). Microorganisms produced by these methods are repeatedly selected and separated for breeding useful microorganisms, so that a mutant strain having properties of interest can be obtained. [0030] Examples of such a mutant strain as used herein include strains obtained from the CM4 strain, the CP3232 strain, or mutant strains thereof as parent strains. Specifically, examples of a mutant strain of the CM4 strain further include mutant strains selected after several repetitive mutations of the CM4 strain. [0031] For example, mutant strains from the Lactobacillus helveticus CM4 strain, the Lactobacillus helveticus CP3232 strain, and mutant strains thereof can be easily distinguished from other lactic acid bacterial strains based on the molecular weight distribution of fragments (of the genomic DNA of a lactic acid bacterium) amplified by the polymerase chain reaction (PCR) method. Briefly, DNA sample of a lactic acid bacterial strain of interest is prepared, gene amplification is performed by the PCR method using primers having the sequences shown in SEQ ID NOS: 1 and 2, for example, the electrophoretic pattern of the thus obtained fragments is analyzed, and whether or not the resultant strains are mutant strains originating from the Lactobacillus helveticus CP3232 strain or the Lactobacillus helveticus CM4 strain can be determined. However, a method for confirming whether or not a subject strain is such a mutant strain is not limited to this method. Even when the above PCR fragments are not detected, a resultant strain may be a mutant strain from the Lactobacillus helveticus CM4 strain or a mutant strain from the Lactobacillus helveticus CP3232 strain. Whether or not a subject strain is such a mutant strain can be confirmed by detection of other gene fragments, microbiologic properties, or techniques known in the art. [0032] Whether or not such a mutant strain has the properties (a) to (d) above, a bacterial strain that can be used as the lactic acid bacterium of the present invention can be obtained. [0033] The lactic acid bacterium of the present invention can be prepared via culture under adequate conditions using a medium generally used for culture of lactic acid bacteria. A natural -11 medium or a synthetic medium can be used as a culture medium as long as it contains carbon sources, nitrogen sources, mineral salts, and other components and it enables culture of lactic acid bacteria with efficiency. Those skilled in the art can adequately select a known medium appropriate for a bacterial strain to be used. Examples of carbon sources that can be used include lactose, glucose, galactose, and blackstrap molasses. Examples of nitrogen sources that can be used include organic nitrogen-containing substances such as casein hydrolysate, whey protein hydrolysate, and soy protein hydrolysate. Examples of mineral salts that can be used include phosphate, sodium, potassium, and magnesium. Examples of an appropriate medium for culture of lactic acid bacteria include MRS liquid medium, GAM medium, BL medium, Briggs Liver Broth, animal milk, skim milk, and milk-derived whey. Preferably, a milk medium containing sterilized powdered skim milk is used. [0034] In addition, culture of the lactic acid bacterium of the present invention can be performed at 20*C to 50*C, preferably 25 0 C to 42*C, and more preferably 28*C to 37 0 C under anaerobic conditions. Temperature conditions can be adjusted using a thermostatic bath, a mantle heater, a jacket, or the like. In addition, the term "anaerobic conditions" used herein refers to a low-oxygen environment in which lactic acid bacteria can proliferate. For instance, anaerobic conditions can be provided by using an anaerobic chamber, an anaerobic box, a sealed container or bag containing a deoxidizer, or the like, or by simply sealing a culture container. The format of culture includes static culture, shake culture, and tank culture. The culture time can be 3-96 hours. It is preferable to maintain the pH of the medium at 4.0-8.0 at the initiation of culture. [0035] A specific example of preparation of the lactic acid bacterium of the present invention is briefly described below. For instance, when Lactobacillus helveticus CP3232 strain is used, the lactic acid bacterium is inoculated to a sterilized milk medium (e.g., a milk medium containing 9.00% (w/w) reconstituted powdered skim milk), followed by overnight culture at 28-37 0 C (for about 18-28 hours). This culture procedure is preferably performed repeatedly. [0036] After culture, the obtained culture product of the lactic acid bacterium may be directly used as is, or may be further optionally subjected to crude purification via centrifugation or the like and/or to solid-liquid separation via filtration or the like and sterilization. [0037] Furthermore, the treated product of a lactic acid bacterium obtained by treating the lactic acid bacterium of the present invention may be used as long as it has properties of interest. -12 - Alternatively, the treated product of a lactic acid bacterium may be subjected to further treatment. Examples of such treatment are described below. [0038] Raw milk, skim milk, or soymilk is fermented using lactic acid bacterial cells and/or a treated product thereof, so that a fermented product can be prepared. For instance, lactic acid bacteria or lactic acid bacteria subjected to another treatment are inoculated to raw milk, skim milk, or soymilk, followed by fermentation under conditions (which are substantially equivalent to the above conditions for culturing lactic acid bacteria) known in the art. The obtained fermented product can be directly used, or it may be subjected to another treatment such as filtration, sterilization, dilution, or concentration. [0039] Lactic acid bacterial cells and/or a treated product thereof can be prepared in the form of suspensions or dilutions by suspending or diluting them in an adequate solvent. Examples of a solvent that can be used include water, physiological saline, and phosphate buffered saline (PBS). [0040] Lactic acid bacterial cells and/or a treated product thereof may be sterilized, so that a sterilized product can be prepared. In order to sterilize lactic acid bacterial cells and/or a treated product thereof, for example, a known sterilization treatment, such as filtration sterilization, radiation sterilization, superheat sterilization, or pressure sterilization, can be performed. [0041] Lactic acid bacterial cells and/or a treated product thereof may be subjected to heat treatment, so that a heat-treated product can be prepared. In order to prepare such heat-treated product, high temperature treatment (for example, at 80-150 C) is performed for lactic acid bacterial cells and/or a treated product thereof for about 10-60 minutes (e.g., about 10-20 minutes). [0042] Lactic acid bacterial cells and/or a treated product thereof is disrupted, ground, or pulverized, so that a disrupted product or a cell-free extract can be prepared. For instance, such treatment can be performed by physical disruption (e.g., agitation or filter filtration), enzymatic lysis treatment, chemical treatment, autolysis treatment, or the like. [0043] Lactic acid bacterial cells and/or a treated product thereof is extracted using an adequate aqueous or organic solvent, so that an extract can be obtained. An extraction method is not particularly limited as long as it is an extraction method using an aqueous or organic solvent - 13 as an extraction solvent. However, an example of such a method is a known method such as a method that involves immersing a lactic acid bacterium or a treated product thereof (prepared by another treatment of a lactic acid bacterium) in an aqueous or organic solvent (e.g., water, methanol, or ethanol), and agitating or refluxing it in the solvent. [0044] In. addition, lactic acid bacterial cells and/or a treated product thereof can be processed into the form of a powdery product (powder) or granular product via drying. Examples of specific drying methods include, but are not particularly limited to, spray drying, drum drying, vacuum drying, and lyophilization, which can be used alone or in combination. Upon drying, an excipient that is generally used may be added as necessary. The lactic acid bacterium of the present invention may also be wet bacterial cells or dry bacterial cells. [0045] Examples of the above treatment may be used alone or in combinations where appropriate. In the present invention, such treated product can be used in a manner similar to that for the lactic acid bacterium. [0046] The above-obtained lactic acid bacterium and/or treated products can be used alone or together with other ingredients as compositions for production of amino acids and/or peptides from animal milk. Specifically, the present invention provides a composition for producing amino acids and/or peptides, containing at least one lactic acid bacterium belonging to Lactobacillus helveticus. [0047] The composition of the present invention enables production of 25 umol/ml or more, preferably 30 ptmol/ml or more, of free amino acids by fermenting animal milk. Preferably, 15% or more of free amino acids produced are branched chain amino acids. Alternatively or additionally, the composition of the present invention enables production of 8.5 mg/ml or more, preferably 10 mg/ml or more, of peptides by fermenting animal milk. Alternatively or additionally, the composition of the present invention enables production of a total amount of 260 pg/ml or more, preferably 280 pg/ml, and more preferably 300 pLg/ml or more of XP peptides and/or XPP peptides by fermenting animal milk. Alternatively or additionally, the composition of the present invention produces 6.0 mg/g or more of IPP and VPP peptides, which amount is converted to the amount of VPP, per acidity of fermented milk obtained by fermenting animal milk at 30-34 0 C. [0048] -14- The composition of the present invention comprises the above lactic acid bacterium and/or treated product thereof as an active ingredient. Specifically, the composition may contain 1 type of lactic acid bacterium and/or a treated product thereof, multiple different lactic acid bacteria and/or treated products, or multiple treated products of lactic acid bacteria (treated differently) in combinations. In addition, the composition of the present invention preferably comprises at least lx107 lactic acid bacterial cells/ml. [0049] Furthermore, the composition of the present invention may be supplemented with, in addition to a lactic acid bacterium as an active ingredient, an additive and an excipient known in the art alone or in combination, as long as they do not inhibit effects of interest. The composition of the present invention may also comprise an additive (e.g., glutamib acid and sugar such as glucose) that accelerates lactic acid bacterial fermentation. The form of the composition of the present invention is not particularly limited, and it may be formulated into a suspension, granules, powders, capsules, or the like. The content of the active ingredient (a lactic acid bacterium) in the composition of the present invention differs depending on the form, generally ranging from 0.0001% by mass to 99% by mass, preferably from 0.001% by mass to 80% by mass, more preferably from 0.001% by mass to 75% by mass, as the amount of lactic acid bacterium. About 107 cells/g to about 1012 cells/g of lactic acid bacterium are contained in the composition of the present invention. [0050] By fermenting animal milk using the lactic acid bacterium or the composition of the present invention, a fermented milk composition containing amino acids and/or peptides at high concentrations is obtained. Therefore, the present invention provides fermented milk that is obtained by fermenting animal milk using at least one lactic acid bacterium belonging to Lactobacillus helveticus, and contains at least one of the following ingredients (a) to (d): (a) 25 pmol/ml or more, preferably 30 kmol/ml, of free amino acids (preferably, 15% or more of the free amino acids are branched chain amino acids.); (b) 8.5 mg/mI or more, preferably 10 mg/mi or more, of peptides; (c) a total amount of 260 Lg/ml or more, preferably 280 [g/ml or more, more preferably 300 ptg/ml or more, of XP peptides and/or XPP peptides; and (d) 6.0 mg/g or more of IPP and VPP peptides, when converted to the amount of VPP, per acidity of fermented milk. [0051] The fermented milk of the present invention can be prepared through fermentation according to a method known in the art using the lactic acid bacterium or the composition of the present -15 invention. Fermentation can be performed by adding at least one lactic acid bacterium to animal milk and then culturing it under appropriate conditions. As animal milk, mammal-derived milk such as cow milk, goat milk, and horse milk, or processed milk such as skim milk, reconstituted milk, or condensed milk, can be used. Specifically, 1 type of milk can be used or multiple types of milk and processed milk can be used in combination. Milk solid content concentration is not particularly limited. For example, nonfat milk solid content concentration when skim milk is used ranges from about 3% by mass to 15% by mass, preferably from 6% by mass to 15% by mass. Animal milk may be sterilized before fermentation. An additive (e.g., glutamic acid or sugar such as glucose) for accelerating lactic acid bacterial fermentation may be added to animal milk. Alternatively, a milk protein ingredient is isolated from animal milk, and then fermentation can also be performed using such milk protein ingredient as a substrate. [0052] As a lactic acid bacterium to be added to animal milk, a pre-cultured lactic acid bacterium is preferably used as a starter. Also, the amount of the lactic acid bacterium to be added is not limited and generally ranges from, in terms of dried lactic acid bacterial cells, 0.005% by mass to 10% by mass, preferably from 0.05% by mass to 5% by mass. Lactic acid bacterium to be used herein may be an intact cultured lactic acid bacterium, a lactic acid bacterium separated from medium via filtration or centrifugation, or a frozen or freeze-dried lactic acid bacterium stored after separation from medium. [0053] Fermentation conditions are almost the same as those for culturing lactic acid bacteria as described above. Fermentation is performed at 20*C to 50*C, preferably 25"C to 42"C, and more preferably 284C to 37*C under anaerobic conditions. In addition, in Examples described later, the lactic acid bacterium of the present invention produced branched chain amino acids and functional peptides in particularly high amounts at 30 0 C to 34 0 C (about 32 0 C). Hence, fermentation is particularly preferably performed at 30'C to 34 0 C. Temperature conditions can be adjusted using a thermostat, a mantle heater, a jacket, or the like. Fermentation can be performed in a form such as static culture, shake culture, or tank culture. Furthermore, the fermentation time can range from 3 hours to 96 hours, and preferably 12 to 36 hours. Fermentation is preferably performed while measuring pH and acidity in a fermenter and keeping pH at 4.0 to 8.0. [0054] Fermented milk obtained as described above may be treated or a treated product of fermented milk may be further treated, as long as it contains desired ingredients. Examples of such -16treatment are as described below. [0055] Fermented milk and/or a treated product thereof is suspended or diluted in an appropriate solvent, so that a suspension or a diluted solution can be prepared. Examples of a solvent that can be used include water, saline, and phosphate buffered saline (PBS). [0056] Fermented milk and/or a treated product thereof is sterilized, so that a sterilized product can be prepared. Fermented milk and/or a treated product thereof can be sterilized by known sterilization methods such as filter sterilization, radiation sterilization, overheat sterilization, and pressure sterilization. [0057] Fermented milk and/or a treated product thereof is subjected to heat treatment, so that a heat-treated product can be prepared. To prepare such heat-treated product, high-temperature treatment (e.g., 80*C to 150*C) is performed for a given time such as about 10-60 minutes (e.g., about 10-20 minutes). [0058] Moreover, fermented milk and/or a treated product thereof is filtered or centrifuged, so that the supernatant (whey) thereof can be prepared. [0059] Furthermore, fermented milk and/or a treated product thereof is dried, so as to obtain a powdery product (powder) or a granular product. Examples of a specific dry method include, but are not particularly limited to, spray drying, drum drying, vacuum drying, and freeze drying. Such a dry method may be employed alone, or these methods may be employed in combination. At this time, an excipient that is generally used may also be added as necessary. [0060] The above examples of treatment may be used alone or in combinations where appropriate. The present invention also encompasses such a treated product of fermented milk. [0061] Fermented milk or a treated product thereof of the present invention contains various amino acids and/or peptides at high concentrations, so that it can be used for applications using the properties of the amino acids and/or peptides. For example, amino acids are involved in flavor, an energy source, a tranquilizing effect, a growth-promoting effect, and the like, so that the fermented milk of the present invention can be used as a flavor improving agent, a nutritional supplement, or an energy supplement. Furthermore, branched chain amino acids - 17 - (BCAAs) including valine, leucine, and isoleucine are essential amino acids and can be used for increasing muscle strength or preventing physical exhaustion. Furthermore, it is considered that the degradation reaction in the liver proceeds because of the presence of alanine (Ala). Furthermore, it is known that Ala induces the taste sensation of umami (good taste and flavor) and has shionare and sunare effects (effects of lowering the sensations of salty and sour tastes) and an effect of masking acrid taste. Lysine (Lys) is said to be the amino acid that is the most easily deficient among the essential amino acids. Lys should be ingested as a raw material such as an antibody, a hormone, an enzyme, or the like. It has also been reported that Lys is also involved in calcium absorption, collagen formation, and bone tissue production. Serine (Ser) is important as a precursor for various in vivo important substances such as phosphatidyl serine and other amino acids. It is also known that Ser suppresses decreases in blood cholesterol level and brain cell death through neuron activation in the brain, or the like, thereby preventing brain aging and serving as a skin moisturizing ingredient. Therefore, the fermented milk or a treated product thereof of the present invention can be used as a flavor improving agent, a nutritional supplement, an energy supplement, a brain function improving agent, an autonomic nerve regulating agent, a parasympathetic nerve regulating agent, a sympathetic nerve regulating agent, an agent for suppressing muscle degradation and muscle damage, a muscle booster, an agent for alleviating fatigue, an endurance improving agent, an alcohol metabolizing agent, or the like. Furthermore, the VPP peptide and IPP peptide have ACE inhibitory activity and a relaxing effect. Specifically, the LPP peptide has ACE inhibitory activity, the YP peptide has a hypotensive effect and an antianxiety effect, and the VP, IP, AP, EP, RP, QP, TP, MP, and GP peptides are non-biodegradable and have ACE inhibitory activity. Therefore, the fermented milk or a treated product thereof of the present invention can be used as a hypotensive drug or a stress relieving agent. [0062] Various amino acid and/or peptide ingredients, or fractions containing these ingredients may be purified from the thus obtained fermented milk or a treated product thereof using a known separation and/or purification method. [0063] The fermented milk or a treated product thereof of the present invention contains at least one ingredient (e.g., amino acids and peptides) from among the ingredients (a) to (d) above, or may contain multiple ingredients in combination. Also the fermented milk or a treated product thereof may contain a lactic acid bacterium used for fermentation. [0064] -18 - Also, the fermented milk of the present invention may be supplemented with an additive, another known hypotensive drug, a stress relieving agent, a nutritional supplement, an energy supplement, a muscle booster, a flavor improving agent, or the like, as described later, alone or in combination, as long as the effect of interest is not inhibited. [0065] The dosage form of the fermented milk of the present invention includes, but not particularly limited to, suspensions (intact fermented milk obtained after fermentation) and tablets, capsules, granules, powders, dust formulations, syrups, and dry syrups treated and obtained by techniques known in the art. The fermented milk is preferably in a dosage form that facilitates oral administration or intake. In addition, a liquid formulation such as a suspension may be suspended in water or another adequate medium immediately before administration or intake. When the fermented milk is formed into tablets or granules, the surfaces may be coated by a known method. [0066] The fennented milk in the above dosage form can be produced according to a conventional method by incorporating generally used additives such as excipients, disintegrators, binders, wetting agents, stabilizers, buffering agents, lubricants, preservatives, surfactants, sweeteners, flavoring agents, aromatics, acidulants, and coloring agents. For example, in a case in which the fermented milk is used for medicines or promotion of health, a pharmaceutically acceptable carrier or an additive can be incorporated into the fermented milk obtained by fermentation. Examples of such pharmaceutically acceptable carriers and additives include water, pharmaceutically acceptable organic solvents, collagen, polyvinyl alcohol, polyvinyl pyrrolidone, carboxyvinyl polymers, sodium alginate, water-soluble dextran, water-soluble dextrin, carboxymethyl starch sodium, pectin, xanthan gum, arabic gum, casein, gelatin, agar, glycerin, propylene glycol, polyethylene glycol, vaseline, paraffin, stearyl alcohol, stearic acid, human serum albumin, mannitol, sorbitol, lactose, and surfactants acceptable as pharmaceutical additives. [0067] Further, the fermented milk of the present invention may further contain a variety of additives used for production of medicines, food or drink products, or feeds and other various substances. Examples of such substances and additives include a variety of fats and oils (e.g., plant oils such as soybean oil, corn oil, safflower oil, and olive oil, and animal fat and oil such as beef fat or sardine oil), herbal medicines (e.g., royal jelly and ginseng), amino acids (e.g., glutamine, cysteine, leucine, and arginine), polyalcohols (e.g., ethylene glycol, polyethylene glycol, propylene glycol, glycerin, and sugar alcohols such as sorbitol, erythritol, - 19 xylitol, maltitol, and mannitol), natural polymers (e.g., arabic gum, agar, water-soluble corn fibers, gelatin, xanthan gum, casein, gluten or gluten hydrolysate, lecithin, starch, and dextrin), vitamins (e.g., vitamin C and vitamin Bs), minerals (e.g., calcium, magnesium, zinc, and iron), dietary fibers (e.g., mannan, pectin, and hemicellulose), surfactants (e.g., glycerin esters of fatty acid and sorbitan esters of fatty acid), purified water, excipients (e.g., glucose, cornstarch, lactose, and dextrin), stabilizing agents, pH adjusting agents, antioxidants, sweeteners, flavoring agents, acidulants, coloring agents, and aromatics. [0068] Further, in addition to the above active ingredients, a functional ingredient or an additive can be incorporated into the fennented milk of the present invention. Examples thereof include taurine, glutathione, carnitine, creatine, coenzyme Q, glucuronic acid, glucuronolactone, capsicum extract, ginger extract, cacao extract, guarana extract, garcinia extract, theanine, y-aminobutyric acid, capsaicin, capsiate, a variety of organic acids, flavonoids, polyphenols, catechins, xanthine derivatives, indigestible oligosaccharides such as fructooligosaccharide, and polyvinyl pyrrolidone. The amount of such additive to be incorporated can be adequately determined depending on the type of additive and the desirable intake. [0069] Subjects of administration or intake of the fermented milk of the present invention may be vertebrate animals. Specific examples thereof include mammals such as humans, primates (e.g., monkeys and chimpanzees), livestock animals (e.g., cattle, horses, pigs, sheep, and birds), pet animals (e.g., dogs and cats), and experimental animals (e.g., mice and rats). Further, such subjects can be reptiles and birds. Particularly preferable subjects are those expected to ingest amino acids and/or peptides, such as hypertensive patients, humans having stress-induced symptoms, and humans before or after exercise. [0070] The dose of administration or intake of the fermented milk of the present invention differs depending on the age and body weight of a subject, an administration/intake route, the number of doses for administration/intake, and the purpose of administration etc., and can be changed extensively at the discretion of those skilled in the art to achieve a desired effect. The contents of an active ingredient contained in the fermented milk is not particularly limited and can be adequately adjusted in accordance with the degree of ease of production, and the preferable daily dose, for example. The fermented milk of the present invention is sufficiently safe and thus the intake dose can also be increased. The daily dose may be ingested in a single dose, or it may be divided into several doses. In addition, the frequency of administration or intake is not particularly limited, and it can be adequately selected -20 depending on various conditions such as an administration/intake route, the age and body weight of a subject, and desired effects. [0071] The administration/intake route of the fermented milk of the present invention is not particularly limited, and oral administration or oral intake is preferred. For example, the fermented milk is incorporated into food or drink products or feedstuff or formulated into tablets, granules, or the like and thus can be orally administered or ingested. [0072] The fermented milk of the present invention may be used in combination with other medicines, treatment methods, prevention methods, or the like. Such other medicines and the fermented milk of the present invention may be formulated into a single formulation. Alternatively, they may be formulated into separate formulations so as to be administered simultaneously or at intervals. [0073] As described above, the fermented milk of the present invention can be used as a hypotensive drug, a brain function improving agent, an autonomic nerve regulating agent, a parasympathetic nerve regulating agent, a sympathetic nerve regulating agent, a nutritional supplement, an energy supplement, an agent for suppressing muscle degradation and muscle damage, a muscle booster, an agent for alleviating fatigue, an endurance improving agent, a flavor improving agent, an alcohol metabolizing agent, or the like. [0074] The lactic acid bacterium and fermented milk of the present invention are sufficiently safe and thus are easily used for long-term continuous intake. Therefore, the lactic acid bacterium and fermented milk of the present invention can also be used for food or drink products and feedstuff. The lactic acid bacterium of the present invention produces a large amount of amino acids and/or peptides having various functions. Moreover, the fermented milk of the present invention contains such amino acids and/or peptides at high concentrations. Furthermore, the lactic acid bacterium of the present invention exhibits the high production amount of functional peptides in relative to acidity, can suppress sourness, and do not inhibit food or drink products' own flavor even when added to various food or drink products. Therefore, the lactic acid bacterium and fennented milk of the present invention are added to various food or drink products, can be continuously ingested, and thus are expected to exhibit various effects. [0075] Therefore, the food or drink product of the present invention contains at least one lactic acid -21 bacterium of the present invention and/or a treated product thereof, and/or the fermented milk of the present invention and/or a treated product thereof. In the present invention, examples of the food or drink product of the present invention also include beverages. Examples of the food or drink product of the present invention include all food or drink products, into which the lactic acid bacterium and fermented milk of the present invention can be incorporated, in addition to health food or drink products for health promotion, functional food or drink products, and food or drink products for specified health use with the use of amino acids and/or peptides. [0076] Functional food or drink products are particularly preferable as the food or drink products of the present invention. The term "functional food or drink product" of the present invention refers to a food or drink product having predetermined functionality for organisms and encompasses, for example, so-called general health food or drink products such as food or drink products with health claims including food or drink products for specified health use (including qualified FOSHU [i.e., food for specified health use]) and food or drink products with nutrient function claims, food or drink products for special dietary uses, nutritional supplements, health supplements, supplements (e.g., those having a variety of dosage forms such as tablets, coated tablets, sugar-coated tablets, capsules, and liquid agents), and beauty food or drink products (e.g., diet food or drink products). [0077] Specific examples of food or drink products include health food or drink products and nutritional supplements in preparation forms such as liquid diets (e.g., tube enteral nutritional supplements), tablet candies, tablets, chewable tablets, tablets, dust formulations, powders, capsules, granules, and tonic drinks; tea beverages such as green tea, oolong tea, and black tea; drinks or beverages such as soft .drinks, jelly beverages, isotonic beverages, milk beverages, carbonated beverages, vegetable beverages, juice beverages, fermented vegetable beverages, fermented juice beverages, fermented milk beverages, fermented milk (e.g., drink yogurt and solid yogurt), fermented milk beverages (sterilized), lactobacillus beverages, condensed beverages, condensed solid materials, milk beverages (e.g., coffee milk), beverages containing drink powders, cocoa beverages, milk, and purified water; spreads such as butter, jam, dried seasoning products, and margarine; mayonnaise; shortening; custard cream; dressings; bread; boiled rice; noodles; pasta; miso soup; tofu; yogurt; soup or sauces; and sweets (e.g., biscuits and cookies, chocolate, candies, cake, ice cream, chewing gum, and tablets). [0078] -22 - The food or drink product of the present invention can be produced according to a conventional method by incorporating other food materials used for production of the above food or drink products, various nutrients, various vitamins, minerals, dietary fibers, and various additives (e.g., taste components, sweeteners, acidulants such as organic acids, stabilizers, and flavors), in addition to the lactic acid bacteria and/or fermented milk of the present invention. [0079] For the food or drink product of the present invention, those skilled in the art can adequately determine the amount of the lactic acid bacterium, fermented milk, or treated product thereof to be incorporated in consideration of the form of the food or drink product and the taste or texture that are required. Through consumption of the food or drink product of the present invention in a form that allows management of the desirable intake thereof, a method using the food or drink product for lowering blood pressure, relieving stresses, nutritional support, energy supplement, enhancing muscle, improving brain functions, regulating autonomic nerve, regulating parasympathetic nerve, regulating sympathetic nerve, suppressing muscle degeneration and damage, enhancing muscle, alleviating fatigue, improving endurance, or alcohol metabolism can be provided, for example. [0080] The food or drink product of the present invention can be produced by an arbitrary appropriate method available by those skilled in the art. For example, the lactic acid bacterium and/or fermented milk (treated product) of the present invention can be prepared in a liquid, gel, solid, powder, or granular form and then incorporated into a food or drink product. Alternatively, the lactic acid bacterium and/or fermented milk (treated product) of the present invention may be mixed or dissolved directly into a raw material for a food or drink product. The lactic acid bacterium and/or fermented milk (treated product) of the present invention may be applied to, coated onto, infiltrated into, or sprayed onto a food or drink product. The lactic acid bacterium and/or fermented milk (treated product) of the present invention may be dispersed uniformly or distributed unevenly in a food or drink product. A capsule containing the lactic acid bacterium and/or fermented milk (treated product) of the present invention may be prepared. An edible film, a food coating agent, or the like may be wrapped around the lactic acid bacterium and/or fermented milk (treated product) of the present invention. Alternatively, the lactic acid bacterium and/or fermented milk (treated product) of the present invention may be prepared into a form such as a tablet after the addition of an appropriate excipient and others. The food or drink product of the present invention may further be processed. Such a processed product is also encompassed -23 within the scope of the present invention. [0081] In the production of the food or drink product of the present invention, a variety of additives as routinely used in food or drink products may be employed. Examples of the additives include, but are not limited to, color formers (e.g., sodium nitrite), coloring agents (e.g., gardenia pigments and Red 102), flavors (e.g., orange flavors), sweeteners (e.g., stevia and aspartame), preservatives (e.g., sodium acetate and sorbic acid), emulsifiers (e.g., sodium chondroitin sulfate and propylene glycol esters of fatty acid), antioxidants (e.g., disodium EDTA and vitamin C), pH adjusters (e.g., citric acid), chemical seasonings (e.g., sodium inosinate), thickeners (e.g., xanthan gum), swelling agents (e.g.,- calcium carbonate), antifoaming agents (e.g., calcium phosphate), binding agents (e.g., sodium polyphosphate), nutrition-enriching agents (e.g., calcium-enriching agents and vitamin A), and excipients (e.g., water-soluble dextrin). Functional raw materials such as Panax ginseng extracts, Acanthopanax senticosus extracts, eucalyptus extracts, or du zhong tea extracts may further be added. [0082] As described above, the food or drink product of the present invention has various functions resulting from amino acids and/or peptides and sufficient safety, but has no concern about side effects. Moreover, the lactic acid bacterium and/or fermented milk of the present invention can suppress acidity, has good flavor, and does not inhibit the flavor of food or drink products when added to various food or drink products. The obtained food or drink product can be easily used for long-term continuous intake with the expectation of long-term exhibition of various functions. [0083] Further, the lactic acid bacterium and/or fermented milk of the present invention can be incorporated not only into food or drink products for humans but also into feeds for animals such as livestock (e.g., cattle and pigs), racehorses, and pets (e.g., dogs and cats). Feeds are substantially equivalent to food or drink products except that they are given to non-human subjects. Therefore, the above descriptions of food or drink products can be applied mutatis mutandis to feeds. EXAMPLES [0084] The present invention is hereafter described in greater detail with reference to the following examples, but the present invention is not limited to the examples. [0085] -24 - [Example 1] Identification of the L. helveticus CP3232 strain The L. helveticus CM4 strain (Accession No. FERM BP-6060) was bred. A novel L. helveticus CP3232 strain that exhibits properties as described in Examples 2 to 5 was obtained from the mutant strains derived from the CM4 strain. [0086] The L. helveticus CP3232 strain was anaerobically cultured in MRS agar medium at 37*C for 24 hours, and then the morphologically observed properties of the obtained colony, the physiological properties thereof, and the like were examined. Furthermore, the 16S rRNA sequence was determined and confirmed to have 99.9% homology with another Lactobacillus helveticus strain (type strain). Therefore, it was revealed that the thus obtained bacterial strain definitely belonged to L. helveticus and was a novel bacterial strain. [0087] [Example 2] Preparation of fermented milk 9.00% (w/w) reconstituted powdered skim milk was sterilized by increasing the temperature to 95*C, and then cooled to 15*C. The resultant was used as a milk medium. Fennented milk containing a lactic acid bacterium (5%) was added (for inoculation) to the milk medium, followed by 24 hours of culture at 37*C. This procedure was repeated, and then the thus obtained fermented milk was used as a starter. The starter (3%) was added to the milk medium and then cultured at 32*C or 37*C for 24 hours. The pH and acidity of the fermented milk were measured. Acidity was measured using the Hiranuma automatic titrator COMTITE-450 (Hiranuma Sangyo Co., Ltd.). As the lactic acid bacteria, L. helveticus CP3232 strain, and for comparison, L. helveticus CP3264 strain (separated from a commercially available antihypertensive product), L. helveticus CM4 strain (Accession No. FERM BP-6060), L. helveticus CP1092 strain, L. helveticus CP1100 strain, L. helveticus CP1 081 strain, and L. helveticus CPN4 strain (JP Patent Publication (Kokai) No. H7-123977 A (1995)), were used. [0088] The results are shown in Table 1. Because ACE inhibitory activity per weight of IPP was 1.7 times that of VPP, the amount of IPP and VPP was calculated in terms of the amount of VPP by the following formula: Amount calculated in terms of amount of VPP (pag/ml) = amount of IPP (p g/ml) x 1.7 + amount of VPP (pig/ml) [0089] In addition, the term "acidity" refers to the concentration of acid (lactic acid) contained in fermented milk. As shown in Table 1, in the case of fermented milk obtained by fermenting -25 milk with the L. helveticus CP3232 strain, the amount of the IPP and VPP peptides in relative to the amount of the produced lactic acid (that is, the acidity) was 6.2 mg/g when converted to the amount of VPP, indicating that the amount of the peptides per acidity was high. Calculation was performed with 1.02 as the specific gravity of fermented milk. [0090] Table I Amount of LTP (IPP and VPP) per acidity (Amount converted to amount of VPP) 320C 370C 24 hours 24 hours (pg/ml) (%) (mg/g) (pg/ml) (%) (mg/g) LTP LTP (Amount(Aon ount Acidity of LTP/ (Amount Acidity of LTP/ cneetoconverted toAcdtof L/ lactic acid Lactic acid lactic acid Lactic acid amount of amount of VPP) VPP) CM4 60.9 1.93 3.1 44.3 2.34 1.9 CP3264 93.9 1.78 5.2 76.8 2.11 3.6 CP3232 96.8 1.53 6.2 76.0 1.94 3.8 CP1092 3.0 0.62 0.5 13.3 1.39 0.9 CP1100 5.8 1.35 0.4 12.0 1.90 0.6 CP1081 71.9 1.65 4.3 72.4 1.95 3.6 CPN4 18.6 1.58 1.2 18.2 1.87 1.0 [0091] [Example 3] Measurement of free amino acids in fermented milk supernatant Fermented milk obtained in Example 2 was subjected to 10 minutes of centrifugation at 10000g. The supernatant was appropriately diluted with distilled water, the solution was filtered with a 0.2 p.M membrane filter, and then measurement was performed by high performance liquid chromatography (HPLC, Shimadzu Corporation) using an amino acid automatic analyzer (C-R7A/LC-10A). Analytical conditions are as described below. [0092] Column: Shim-pack Amino-Li (100 mmL x 6.0 mmI.D.) Ammonia trap: Shim-pack ISC-3OLi (50 mmL x 4.0 mmI.D.) Column temperature: 384C to 58*C Reaction temperature: 65 0 C Mobile phase: Shimadzu mobile phase kit for amino acid analysis, i.e., the type Li Reaction solution: Shimadzu reaction solution kit for amino acid analysis, i.e., the OPA -26 reagent Flow rate: 0.4 ml/min. [0093] Sodium hypochlorite was added to the reaction solution for proline detection. Fluorescence color development of amino acids that had finally reacted with o-phthalaldehyde was measured with a spectrofluoro-detector (Ex: 348 nm; Em: 450 nm). A standard amino acid mixture (Type H), which has a known concentration of each amino acid, was used as the standard sample, and then the contents of amino acids were calculated. [0094] The results are shown in Fig. 1 and Fig. 2. Fig. 1A shows the results of measuring the amount of each amino acid (nrol/ml) contained in the supernatant of the fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 32*C for 24 hours. Fig. 1B shows the results obtained by measuring the amount of each amino acid (nmo1/ml) contained in the supernatant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 37*C for 24 hours. Fig. 2A shows the total amount of free amino acids (ptmol/ml) contained in the supernatant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 32*C or 37"C for 24 hours. Fig. 2B shows the amount of branched chain amino acids (pmol/ml) contained in the supernantant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 32*C or 37*C for 24 hours. It was demonstrated from these results that the L. helveticus CP3232 strain has a high proteolysis activity and produces 25 mol/ml or more of free amino acids by 24 hours of fermentation. In particular, the L. helveticus CP3232 strain produced 29.9 ptmol/ml of free amino acids by fermenting milk at 37*C, while the same strain produced 33.1 pmol/ml of free amino acids by fermenting milk at 32*C (Fig. 2A). It was also demonstrated that the amount of branched chain amino acids contained in the produced free amino acids was high, and, 15% or more of all free amino acids produced by 24 hours of fermentation were branched chain amino acids. Specifically, branched chain amino acids represented about 19.4% of all free amino acids in the case of fermentation at 37"C, and branched chain amino acids represented about 21.5% of all free amino acids in the case of fermentation at 32*C (Fig. 2B). [0095] [Example 4] Amount of peptides in fermented milk supernatant (OPA method) The amounts of peptides in the fermented milk supernatants obtained in Example 2 were measured according to the method of Church et al., (J. Dairy Sci. 1983. 66: 1219-1227). Briefly, 25 ml of 100 mM sodium tetraborate solution, 2.5 ml of 20% SDS, and 0.1 ml of -27 - 2-mercaptoethanol were mixed. One (1) ml of a solution prepared by dissolving o-phthalaldehyde in methanol (4% by weight) was added to the mixture, and then the solution was finally adjusted with distilled water to 50 ml, thereby preparing an OPA reagent. Fermented milk obtained in Example 2 was subjected to centrifuging it at 10,000g for 10 minutes, and the obtained culture supernatant was appropriately diluted, so as to prepare an analysis sample. The OPA reagent (200 pl) and the analysis sample (10 pl) were mixed. After sufficient stirring, the mixture was left to stand at room temperature for 5 minutes, and then absorption at 340 mn was measured. As a reference material, bacto tryptone was used. [0096] The results are shown in Fig. 3. It was demonstrated from the results that the L. helveticus CP3232 strain has a high proteolysis activity and produces a large amount of peptides through fermentation. Specifically, the L. helveticus CP3232 strain produced 8.5 mg/ml or more of peptides through 24 hours of fennentation, 11.1 mg/ml of peptides through fermentation at 32*C, and 9.6 mg/ml of peptides through fermentation at 37 0 C (Fig. 3). [0097] [Example 5] Quantification of functional peptides Fermented milk obtained in Example 2 was subjected to 10 minutes of centrifugation at 10,000g, and the supernatant was appropriately diluted with distilled water and then filtered with 0.2 1 4M membrane filter before analysis on LC/MS. LC/MS analysis was performed by the method of Inoue et al. (J. Biosci. Bioeng., 2009, 108, 111-115). Specifically, the analysis was conducted using the LCMS-2010A system (Shimadzu Corporation) and the RP-aqueous column (Nomura Chemical Co., Ltd.) as a separation column. Contents were calculated using synthetic peptides as the standard samples. The synthetic peptides used herein were (1 3 C 5 )Val-(' 3 C)Pro-Pro, Ile-(' 3
C
5 )Pro-Pro, Glu-Pro, and Gln-Pro (purchased from Scrum Inc.), and Leu-Pro-Pro, Tyr-Pro, Val-Pro, Ile-Pro, Ala-Pro, Arg-Pro, Thr-Pro, Met-Pro, and Gly-Pro (purchased from Bachem). [0098] Fig. 4 and Fig. 5 show the results. It was revealed by the results shown in Fig. 4 A and Fig. 4B that the L. helveticus CP3232 strain has a high proteolysis activity and produces various functional peptides through 24 hours of fermentation. It has been reported that among functional peptides listed in Fig. 4, VPP and IPP have ACE inhibitory activity and relaxing effects, LPP has ACE inhibitory activity, YP has a hypotensive effect and an anti-anxiety effect, and VP, IP, AP, EP, RP, QP, TP, MP, and GP are non-biodegradable and have ACE inhibitory activity. The L. helveticus CP3232 strain produced particularly large amounts of VPP, IPP, YP, QP, and TP. -28 - [0099] It was further revealed by the results shown in Fig. 5A and Fig. 5B that the L. helveticus CP3232 strain has a high proteolysis activity and produces a total of 260 pg/ml or more of functional peptides (XP + XPP) through 24 hours of fermentation. In particular, the L. helveticus CP3232 strain produced 318 pmol/ml of functional peptides (Fig. 5A) through fermentation at 32*C and 282 pmol/ml of functional peptides through fermentation at 37*C, respectively (Fig. SB). [0100] [Example 6] Preparation of fermented milk and measurement of amount of peptides In a manner similar to Example 2, the pH and acidity of fermented milk and the amount of the thus produced IPP peptide and VPP peptide were measured after culture using, as lactic acid bacteria, the L. helveticus CP3232 strain, and for comparison, the L. helveticus CNCM 1-3435 strain (Patent Literature 4), the L. helveticus FERM BP-5445 strain (Patent Literature 7), and the L. helveticus CM4 strain (Accession No. FERM BP-6060). [0101] The results are shown in Table 2. The results are expressed in a manner similar to that in Example 2; that is, the amount of IPP and VPP was indicated by being converted to the amount of VPP. The term "acidity" refers to the concentration of acid (lactic acid) contained in fermented milk. As shown in Table 2, in the case of fermented milk obtained by fermentation with the L. helveticus CP3232 strain, the amount of IPP and VPP peptides in rlative to lactic acid (that is, the acidity) produced was 7.0 mg/g when converted to the amount of VPP, indicating that the amount of the peptides per unit of acidity was high. Calculation was performed with 1.02 as the specific gravity of fermented milk. [0102] [Table 2] -29- Amount of LTP (IPP and VPP) per acidity (Amount converted to amount of VPP) 32*C 37"C 24 hours 24 hours (pg/ml) (%) (mg/g) (pg/ml) (%) (mg/g) LTP LTP (Amount(Aon ount Acidity of LTP/ (Amount Acidity of LTP/ cneeto .converted toAcdtof LP lactic acid Lactic acid lactic acid Lactic acid amount of amount of VPP) VPP) CM4 61.1 1.92 3.1 44.3 2.29 1.9 CP3232 96.2 1.35 7.0 79.0 1.99 3.9 1-3435 86.1 1.79 4.7 79.9 2.10 3.7 BP-5445 18.5 1.12 1.6 26.9 1.40 1.9 [0103] [Example 7] Measurement of free amino acids in fermented milk supernatant Free amino acids were measured in a manner similar to that in Example 3 using fermented milk obtained in Example 6. The results are shown in Fig. 6 and Fig. 7. [0104] Fig. 6A shows the amount of each amino acid (nmol/ml) contained in the supernatant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 32*C for 24 hours. Fig. 6B shows the amount of each amino acid (nmol/ml) contained in the supernatant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 37*C for 24 hours. Also, Fig. 7A shows the total amount of free amino acids (p mol/ml) contained in the supernatant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 32*C or 37*C for 24 hours. Fig. 7B shows the amount of branched chain amino acids (pLmol/ml) contained in the supernatant of fermented milk obtained by fermenting milk with each of the indicated lactic acid bacterial strains at 320C or 37*C for 24 hours. It was demonstrated from these results that the L. helveticus CP3232 strain has a high proteolysis activity and produces 25 mol/ml or more of free amino acids through 24 hours of fermentation. In particular, the L. helveticus CP3232 strain produced 28.1 [imol/ml of free amino acids through fermentation at 370C and 32.4 gmol/ml thereof through fermentation at 32*C (Fig. 7A). It was also demonstrated that the amount of branched chain amino acids contained in the thus produced free amino acids was high and branched chain amino acids represented 15% or more of free amino acids produced through 24 hours of fermentation. Specifically, branched chain amino acids represented about 20% of free amino acids in the case of -30fermentation at 37*C. Branched chain amino acids represented about 21% of free amino acids in the case of fermentation at 32 0 C (Fig. 7B). [0105] [Example 8] Amount of peptides in fermented milk supernatant (OPA method) The amounts of peptides in fermented milk supernatants obtained in Example 6 were measured in a manner similar to that in Example 4. The results are shown in Fig. 8. It was demonstrated by the results that the L. helveticus CP3232 strain has a high proteolysis activity and produces a large amount of peptides through fermentation. Specifically, the L. helveticus CP3232 strain produced 8.5 mg/mil or more of peptides through 24 hours of fermentation, 10.4 mg/ml of peptides through fermentation at 32*C, and 9.1 mg/ml of peptides through fennentation at 37*C (Fig. 8). [0106] [Example 9] Quantification of functional peptides Fermented milk obtained in Example 6 was analyzed by LC/MS in a manner similar to that in Example 5, and then the amounts of functional peptides were measured. Fig. 9 and Fig. 10 show the results. It was revealed by the results shown in Fig. 9A and Fig. 9B that the L. helveticus CP3232 strain has a high proteolysis activity and produces various functional peptides through 24 hours of fermentation. The L. helveticus CP3232 strain produced larger amounts of VPP, IPP, YP, QP, and TP in particular. [0107] Furthermore, it was demonstrated from the results shown in Fig. 10 A and Fig. 1 0B that the L. helveticus CP3232 strain has a high degree of proteolysis activity and produces 260 jig/ml or more (in total) of functional peptides (XP+XPP) through 24 hours of fermentation. In particular, the L. helveticus CP3232 strain produced 280 pmol/ml (Fig. 10A) of functional peptides through fermentation at 32*C and 295 yimol/ml of functional peptides through fermentation at 37*C (Fig. 1GB). [0108] [Reference Example 1] Method for identifying L. helveticus CP3232 strain and mutant strain thereof Lactic acid bacteria, i.e., L. helveticus CP3232 strain, and mutant strains thereof, which weer obtained in Example 1, can be easily distinguished from other lactic acid bacterial strains by combining measurement by the methods described in Examples 2 to 9 with identification by the polymerase chain reaction (PCR) method. Furthermore, whether or not a strain is a mutant strain of the L. helveticus CM4 strain, which is the parent strain of the L. helveticus CP3232 strain, can be conveniently detennined by the PCR method. -31- [0109] One example is briefly described below. Whether or not a lactic acid bacterial strain of interest is a mutant strain of the L. helveticus CP3232 strain or a mutant strain of the L. helveticus CM4 strain can be determined by preparing a DNA sample, performing gene amplification by the PCR method using the following primers, performing electrophoresis with 0.9% agarose gel, and then analyzing the electrophoretic pattern. [0110] Primers: agccacttcctccgattacag (SEQ ID NO: 1); and gctattttagcagcgattcg (SEQ ID NO: 2). [0111] In this Example, the above PCR method was performed using multiple L. helveticus strains. Fig. 11 shows the results. In Fig. 11, each lane is as follows. [0112] Lane 1: L. helveticus CP3232 strain Lane 2: L. helveticus CM4 strain Lane 3: L. helveticus CP3231 strain Lane 4: L. helveticus CP3264 strain Lane 5: L. helveticus CP 1081 strain Lane 6: L. helveticus CP209 strain Lane 7: L. helveticus CP2 10 strain Lane 8: L. helveticus CP293 strain Lane 9: L. helveticus CP617 strain Lane 10: L. helveticus CP39 strain Lane 11: L. helveticus CP790 strain Lane 12: L. helveticus JCM1 120 strain M: restriction enzyme Hind III digest of Xphage DNA. [0113] As shown in Fig. 11, a characteristic DNA fragment was observed for CM4 mutant strains (the L. helveticus CP3232 strain and CP3231 strain) and the parent CM4 strain (lanes 1 to 3). But the fragment was not observed for other L. helveticus strains. The restriction enzyme Hind III digest of Xphage DNA was simultaneously electrophoresed as a DNA marker and then the molecular weight of a characteristic DNA fragment was determined to be about 2.2 kb. [0114] -32- In addition, the L. helveticus CP3232 strain and CP3231 strain, and the parent CM4 strain were identified as follows. The L. helveticus CP3232 strain can also be selected by analyzing the amount of peptides in fermented milk using the OPA reagent, as described in Examples 4 and 8. Specifically, the amounts of peptides in fermented milk prepared according to Example 2 as measured using the OPA reagent are as follows. Whereas the L. helveticus CP3231 strain and the CM4 strain produced 7.5 mg/ml and 4.5 mg/ml of peptides, respectively, the L. helveticus CP3232 strain produced as much as 11.1 mg/ml of peptides, and thus it could be easily distinguished from other strains. [0115] [Reference Example 2] Method for identification of L. helveticus CP3232 strain and mutant strain thereof In a manner similar to that in Reference example 1, whether or not lactic acid bacteria obtained in Example 1, the L. helveticus CP3232 strain and a mutant strain thereof, can be distinguished from other lactic acid bacterial strains (the L. helveticus CNCM 1-3435 strain (Patent Literature 4) and the L. helveticus FERM BP-5445 strain (Patent Literature 7)) by the polymerase chain reaction (PCR) method was tested. [0116] Fig. 12 shows the results of performing gene amplification by the PCR method using primers and performing electrophoresis. In Fig. 12, each lane is as follows: Lane 1: L. helveticus CP3232 strain Lane 2: L. helveticus CM4 strain Lane 3: L. helveticus CNCM 1-3435 strain Lane 4: L. helveticus FERM BP-5445 strain M: restriction enzyme Hind III digest of Xphage DNA. [0117] As shown in Fig. 12, a characteristic DNA fragment was observed for the L. helveticus CP3232 strain (CM4 mutant strain) and the parent CM4 strain (lanes 1 and 2), but the fragment was not observed for other L. helveticus strains. [0118] All publications, patents, and patent applications cited herein are incorporated herein by reference in their entirety. [Industrial Applicability] [0119] According to the present invention, lactic acid bacteria belonging to Lactobacillus helveticus having high proteolysis activity and good peptide production ability and/or amino acid -33 production ability are provided. Amino acids and/or peptides produced by the lactic acid bacteria have various functions. For example, peptides such as IPP, VPP, and YP have hypotensive effects. Furthermore, with the use of the lactic acid bacteria for fermentation, fermented milk containing various amino acids and/or peptides having such various functions at high concentrations and good flavor can be obtained. Therefore, the present invention is useful in the fields of medicine, food and drink products, health promotion, and the like. [Accession Number] [0120] Accession No: FERM BP-1 1271 (Lactobacillus Helveticus CP3232 strain, deposited on August 4, 2010) Accession No: FERM BP-6060 (Lactobacillus Helveticus CM4 strain, deposited on August 15, 1997) [Sequence Listing Free Text] [0121] SEQ ID NOS: 1 and 2: DNAs (synthetic oligonucleotides) -34-
Claims (18)
1. An isolated lactic acid bacterium belonging to Lactobacillus helveticus, having the following properties: 5 (a) producing 25 imol/ml or more of free amino acids by fermenting animal milk; and (b) producing 8.5 mg/ml or more of peptides by fermenting animal milk; and (c) at least one of: (i) producing XP peptides and XPP peptides in a total amount of 260 pg/ml or more by fermenting animal milk; and 10 (ii) producing 6.0 mg/g or more of IPP and VPP peptides, which amount is converted to the amount of VPP, per acidity of fermented milk that is obtained by fermenting animal milk at 30-34*C.
2. The lactic acid bacterium according to claim 1, wherein 15% or more of the free 15 amino acids are branched chain amino acids.
3. The lactic acid bacterium according to claim 1 or 2, which produces YP peptide in an amount of 60 ptg/ml or more. 20 4. The lactic acid bacterium according to any one of claims 1 to 3, which is a mutant strain of Lactobacillus helveticus CM4 strain (Accession No. FERM BP-6060).
5. The lactic acid bacterium according to any one of claims 1 to 4, which is Lactobacillus helveticus CP3232 strain (Accession No. FERM BP- 11271) or a mutant 25 strain thereof.
6. Lactobacillus helveticus CP3232 strain (Accession No. FERM BP-1 1271).
7. A composition for producing amino acids and/or peptides from animal milk, 30 comprising at least one lactic acid bacterium belonging to Lactobacillus helveticus having the following properties: (a) produces 25 [tmol/ml or more of free amino acids by fermenting animal milk; and (b) produces peptides in an amount of 8.5 mg/ml or more by fermenting animal 35 milk; and (c) at least one of: 35 (i) produces XP peptides and XPP peptides in a total amount of 260 g/ml or more by fermenting animal milk; (ii) produces YP peptide in an amount of 60 pg/ml or more by fermenting animal milk; and 5 (iii) produces 6.0 mg/g or more of IPP and VPP peptides, which amount is converted to the amount of VPP, per acidity of fermented milk that is obtained by fermenting animal milk at 30-34'C.
8. The composition according to claim 7, wherein 15% or more of the free amino acids 10 at (i) are branched chain amino acids.
9. The composition according to any one of claims 7 or 8, wherein the lactic acid bacterium is of any one of claims 1 to 6. 15 10. A culture comprising at least one lactic acid bacterium belonging to Lactobacillus helveticus having the following properties: (a) produces 25 tmol/ml or more of free amino acids by fermenting animal milk; and (b) produces peptides in an amount of 8.5 mg/ml or more by fermenting animal 20 milk; and (c) at least one of: (i) produces XP peptides and XPP peptides in a total amount of 260 tg/ml or more by fermenting animal milk; (ii) produces YP peptide in an amount of 60 [tg/ml or more by fermenting 25 animal milk; and (iii) produces 6.0 mg/g or more of IPP and VPP peptides, which amount is converted to the amount of VPP, per acidity of fermented milk that is obtained by fermenting animal milk at 30-34'C. 30 11. The culture according to claim 10, wherein the culture comprises at least one additional bacterium.
12. Fermented milk, which is obtained by fermenting animal milk with at least one lactic acid bacterium belonging to Lactobacillus helveticus having the following 35 properties: (a) 25 tmol/ml or more of free amino acids; and 36 (b) 8.5 mg/ml or more of peptides; and (c) at least one of: (i) 260 tg/ml or more in total of XP peptides and XPP peptides; and (ii) 6.0 mg/g or more of IPP and VPP peptides per acidity when converted to the 5 amount of VPP.
13. The fermented milk according to claim 12, comprising at least one lactic acid bacterium belonging to Lactobacillus helveticus. 10 14. The fermented milk according to claim 12 or 13, comprising at least one lactic acid bacterium of any one of claims 1 to 6.
15. The fermented milk according to any one of claims 12 to 14, wherein 15% or more of the free amino acids are branched chain amino acids. 15
16. The fermented milk according to any one of claims 12 to 15, comprising 60 pg/ml or more of YP peptide.
17. The fermented milk according to any one of claims 12 to 16, wherein the 20 fermentation is performed at a temperature of 30-34'C.
18. Use of the fermented milk according to any one of claims 12 to 17, as a or in the manufacture of a hypotensive drug, a brain function improving agent, an autonomic nerve regulating agent, a parasympathetic nerve regulating agent, a sympathetic nerve 25 regulating agent, a nutritional supplement, an energy supplement, an agent for suppressing muscle degradation and muscle damage, a muscle booster, an agent for alleviating fatigue, an endurance improving agent, a flavor improving agent, or an alcohol metabolizing agent. 30 19. A food or drink product, comprising at least one lactic acid bacterium of any one of claims 1 to 6 and/or a treated product thereof, and/or the fermented milk of any one of claims 12 to 17 and/or a treated product thereof.
20. A method for producing an amino acid(s) and/or a peptide(s), comprising 35 fermenting animal milk or milk proteins using at least one lactic acid bacterium of any 37 one of claims 1 to 6 or the composition of any one of claims 7 to 8, and then collecting the amino acid(s) and/or the peptide(s) from the obtained fermented product.
21. The method according to claim 20, wherein the amino acid is a branched chain 5 amino acid.
22. The method according to claim 20 or 21, wherein the peptide is at least one peptide selected from the group consisting of XP peptides and/or XPP peptides. 10 32. The method according to any one of claims 20 to 22, wherein the fermentation is performed at a temperature of 30-34'C.
24. An isolated lactic acid bacterium belonging to Lactobacillus helveticus or a composition or a fermented milk or a use or a food or drink product or a method 15 substantially as described herein with reference to any one or more of the Examples and/or Drawings. 38
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CN112029784A (en) * | 2020-09-16 | 2020-12-04 | 内蒙古农业大学 | Gene related to proteolytic ability and screening method of lactobacillus helveticus with high proteolytic ability based on gene |
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EP1016709A1 (en) * | 1997-09-26 | 2000-07-05 | Calpis Co., Ltd. | Lactobacillus helveticus bacterium having high capability of producing tripeptide, fermented milk product, and process for preparing the same |
JP2005517424A (en) * | 2002-02-25 | 2005-06-16 | ヴァリオ・オサケ・ユキテュア | Improving mineral bioavailability with biologically active peptides |
WO2009149880A1 (en) * | 2008-06-12 | 2009-12-17 | Nestec S.A. | Lactobacillus helveticus strains for producing hypotensive peptides |
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US5854029A (en) * | 1996-08-02 | 1998-12-29 | Calpis Co., Ltd. | Method for preparing antihypertensive agent |
WO2006004105A1 (en) * | 2004-07-05 | 2006-01-12 | Calpis Co., Ltd. | Method of producing peptide mixture, method of producing fermented milk containing antihypertensive peptide and method of producing antihypertensive peptide preparation |
PL1820851T3 (en) * | 2006-02-20 | 2012-07-31 | Gervais Danone Sa | Non-lactose fermenting Lactobacillus helveticus |
ATE423840T1 (en) * | 2006-02-20 | 2009-03-15 | Gervais Danone Sa | NEW STRAINS OF LACTOBACILLUS HELVETICUS |
JP2009296972A (en) * | 2008-06-16 | 2009-12-24 | Snow Brand Milk Prod Co Ltd | Enzyme modified cheese, and method for producing the same |
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2011
- 2011-11-08 CN CN201180054138.4A patent/CN103221531B/en not_active Expired - Fee Related
- 2011-11-08 MY MYPI2013700546A patent/MY160992A/en unknown
- 2011-11-08 JP JP2012542934A patent/JP5888701B2/en not_active Expired - Fee Related
- 2011-11-08 WO PCT/JP2011/075735 patent/WO2012063826A1/en active Application Filing
- 2011-11-08 AU AU2011327288A patent/AU2011327288B2/en not_active Ceased
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Patent Citations (3)
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EP1016709A1 (en) * | 1997-09-26 | 2000-07-05 | Calpis Co., Ltd. | Lactobacillus helveticus bacterium having high capability of producing tripeptide, fermented milk product, and process for preparing the same |
JP2005517424A (en) * | 2002-02-25 | 2005-06-16 | ヴァリオ・オサケ・ユキテュア | Improving mineral bioavailability with biologically active peptides |
WO2009149880A1 (en) * | 2008-06-12 | 2009-12-17 | Nestec S.A. | Lactobacillus helveticus strains for producing hypotensive peptides |
Also Published As
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CN103221531A (en) | 2013-07-24 |
AU2011327288A1 (en) | 2013-05-02 |
MY160992A (en) | 2017-03-31 |
CN103221531B (en) | 2018-03-16 |
TW201305331A (en) | 2013-02-01 |
JP5888701B2 (en) | 2016-03-22 |
TWI551684B (en) | 2016-10-01 |
JPWO2012063826A1 (en) | 2014-05-12 |
WO2012063826A1 (en) | 2012-05-18 |
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