U.S. patent application number 13/380152 was filed with the patent office on 2012-04-26 for novel leuconostoc citreum and fermented foods using the same as a starter, and compositions thereof.
Invention is credited to Heon Woong Jung, Bong Joon Kim, Min-Soo Kwon, Kang-Pyo Lee, Ji-Young Oh.
Application Number | 20120100256 13/380152 |
Document ID | / |
Family ID | 43613920 |
Filed Date | 2012-04-26 |
United States Patent
Application |
20120100256 |
Kind Code |
A1 |
Kim; Bong Joon ; et
al. |
April 26, 2012 |
NOVEL LEUCONOSTOC CITREUM AND FERMENTED FOODS USING THE SAME AS A
STARTER, AND COMPOSITIONS THEREOF
Abstract
The present invention relates to Leuconostoc citreum CJGN34 KCTC
10974BP lactic acid bacteria, fermented foods including kimchi
manufactured using the lactic acid bacteria as a starter, and
lactic acid bacteria composition comprising of the lactic acid
bacteria. The novel Leuconostoc citreum CJGN34 of the present
invention enhances the general taste quality of kimchi; increases
the intrinsic complex sour taste and carbonated taste of kimchi
manufactured by traditional manufacturing method in wintertime; and
regularly sustains the fermentation quality during the year;
particularly, has an effect for controlling the deterioration of
fermentation quality in the summer in which the taste quality
deteriorates. Thus, the lactic acid bacteria of the present
invention can be used as a starter in fermented foods including
kimchi, or as a drug medicine in the form of a tablet or capsule
mixed with carriers or additives, or a s probiotics for foods; or
applied in cosmetic ingredients by mixing in a certain amount.
Therefore, the lactic acid bacteria of the present invention can be
used in a manner commonly applied in various fields of technology
such as medicine, food, feed, cosmetics or the like.
Inventors: |
Kim; Bong Joon; (Incheon,
KR) ; Oh; Ji-Young; (Gyeonggi-do, KR) ; Kwon;
Min-Soo; (Gyeonggi-do, KR) ; Jung; Heon Woong;
(Seoul, KR) ; Lee; Kang-Pyo; (Seoul, KR) |
Family ID: |
43613920 |
Appl. No.: |
13/380152 |
Filed: |
September 10, 2009 |
PCT Filed: |
September 10, 2009 |
PCT NO: |
PCT/KR2009/005139 |
371 Date: |
December 22, 2011 |
Current U.S.
Class: |
426/61 ; 426/531;
426/615; 435/252.9 |
Current CPC
Class: |
A61K 8/99 20130101; A23V
2002/00 20130101; A23Y 2260/21 20130101; C12R 1/01 20130101; A23L
33/135 20160801; C12N 1/20 20130101; A23K 10/18 20160501; A61K
2800/10 20130101; A23L 29/065 20160801; A23L 19/20 20160801; A61Q
19/00 20130101 |
Class at
Publication: |
426/61 ; 426/531;
426/615; 435/252.9 |
International
Class: |
A23L 1/23 20060101
A23L001/23; A23L 1/30 20060101 A23L001/30; C12N 1/20 20060101
C12N001/20; A23L 1/218 20060101 A23L001/218 |
Foreign Application Data
Date |
Code |
Application Number |
Jul 16, 2009 |
KR |
10-2009-0065024 |
Aug 19, 2009 |
JP |
2009-190443 |
Claims
1. Leuconostoc citreum CJGN34 KCTC 10974BP.
2. A starter composition for fermenting foods, which contains
Leuconostoc citreum CJGN34 KCTC 10974BP as an effective
ingredient.
3. A fermented food manufactured by using the starter composition
for fermenting foods in claim 2.
4. A pickled food manufactured by using the starter composition for
fermenting foods in claim 2.
5. Kimchi manufactured by using the starter composition for
fermenting foods in claim 2.
6. A pharmaceutical composition of lactic acid bacteria, which
contains Leuconostoc citreum CJGN34 KCTC 10974BP as an effective
ingredient.
7. A food composition of lactic acid bacteria, which contains
Leuconostoc citreum CJGN34 KCTC 10974BP as an effective ingredient.
Description
FIELD OF THE INVENTION
[0001] The present invention relates to Leuconostoc citreum
bacteria, fermented foods using the bacteria as a starter, and
compositions containing the bacteria. More particularly, the
present invention relates to Leuconostoc citreum which can induce
fermentation flavor and taste quality of Kimchi manufactured in
winter throughout a year and, more importantly, can effectively
inhibit deterioration of fermentation quality in summer, fermented
foods using the bacteria as a starter, and compositions containing
the bacteria.
BACKGROUND OF THE INVENTION
[0002] Kimchi is a collective name of fermented vegetable foods
representing Korea, which are prepared by salting various
vegetables, blending with various seasonings and naturally aging.
Especially, representative Kimchi is Gimjang (``) Kimchi, a main
ingredient of which is Korean cabbage, i.e., Brassica campestris L.
Gimjang Kimchi is prepared by salting Korean cabbage and radish
harvested at late autumn to early winter, blending them with
sub-ingredients such as garlic, ginger, green onion and so on and
burying them underground and is served as winter food (Cheigh and
Park, Biochemical, microbiological and nutritional aspects of
Kimchi, Crit. Rev. FoodSci. Nutr. Vol. 34, 1981, pages 102-127).
Like this, Gimjang Kimchi is categorized as cold fermentation
preserves and is manufactured by keeping vegetables at low
temperature during winter season when vegetables are not produced,
through salting and fermentation methods. Gimjang Kimchi is served
as sources of carbon, amino acid and vitamins and provides a rich
fermentation flavor formed from processes of fermentation and aging
with unique lactic acid bacteria in raw materials.
[0003] Recently, however, raw materials such as Korean cabbages can
be produced and stored throughout a year owing to bread improvement
and cultivation development and, therefore, Korean cabbage Kimchi
can be manufactured even though it is not Gimjang season (i.e.,
winter). Further, the spread of refrigerator and Kimchi
refrigerator have enables cold fermentation preservation as if it
were winter. As a result, recent Kimchi is no more the same with
the traditional Korean cabbage Gimjang Kimchi.
[0004] Basically, the quality of Kimchi significantly changes
depending on that of various raw materials such as Korean cabbage
which is recently forwarded throughout a year. The quality change
of Kimchi has become an obstacle to providing products having
constant quality with customers, especially in a position of
business companies commercializing Kimchi. Especially, Kimchi
manufactured in summer (June.about.August) has problems like low
quality of raw materials and deterioration of fermentation-aging
flavor even though the Kimchi would be manufactured by the same
fermentation and aging methods as in winter. It is well known in
the art that a wide variation in quality depending on season has
brought about various claims of quality dissatisfaction by
customers.
[0005] Prior arts for solving the problems have tried to control
Kimchi quality by adding various microorganisms, mainly lactic acid
bacteria, as a starter (Korean Patent Nos. 10-0536108, 10-0181009,
10-0330674 and so on). The prior arts, however, applied starters
merely for improving quality or confirmed that there is a
difference of general taste quality in sensory test, regardless of
season; they did not investigate causes of quality change during a
year. Accordingly, there is no prior art that focuses on the
quality improvement of summer Kimchi, analyzes the causes of
quality deterioration and solves the problems.
[0006] Meanwhile, Kimchi lactic acid bacteria such as Leuconostoc
Kimchii, Leuconostoc citreum, Leuconostoc mensenteroides,
Leuconostoc gasicomitatum, Leuconostoc lactis, Lactobacillus
plantarum and Lactobacillus sakei have been used individually or in
combination in manufacturing various fermented foods.
[0007] Among them, Leuconostoc citreum bacteria is concerned in
Kimchi aging and is known as its ability to produce polymer
mucilages, dextran, with sugar (Korean Patent No. 10-0718344) and
as being effective for treatment and prevention of atopic
dermatitis (Korean Patent Application No. 10-2005-0041572).
Further, Korean Patent No. 10-0814665 discloses Leuconostoc citreum
S5 bacteria producing dextran and a method of producing fermented
products comprising the dextran by using the bacteria and Korean
Patent No. 10-080053 discloses Leuconostoc citreum producing
mannitol and a method of producing mannitol using the bacteria.
SUMMARY OF THE INVENTION
[0008] The present inventors analyzed fermentation patterns of
Korean cabbage Kimchi during a year and monitored the changes of
lactic acid bacteria flora and organic acids which cause changes of
fermentation quality.
[0009] By doing so, the present inventors found out that lactic
acid bacteria isolated from Kimchi manufactured in winter when the
most delicious Kimchi can be manufactured, have a significant role
in improving and stabilizing quality throughout a year. Further,
the present inventors found out that the lactic acid bacteria can
induce the fermentation flavor and taste quality of winter Kimchi
and effectively inhibit deterioration of fermentation quality in
summer.
[0010] Therefore, a purpose of the present invention is to provide
a novel Leuconostoc spp. lactic acid bacteria which can be used as
a starter for Kimchi, induce fermentation flavor and taste quality
of winter Kimchi and effectively inhibit deterioration of
fermentation quality in summer.
[0011] Further, another purpose of the present invention is to
provide fermented foods manufactured by using the novel Leuconostoc
spp. lactic acid bacteria as a starter.
[0012] In addition, another purpose of the present invention is to
provide a bacteria composition containing the novel Leuconostoc
spp. lactic acid bacteria.
[0013] In order to achieve such purposes, the present invention
provides Leuconostoc citreum CJGN 34 (Leucoostoc citreum CJGN34)
(Deposited with Korean Type of Collection Culture (KCTC) on 2006.
Jul. 27, Accession number: KCTC 10974BP).
[0014] Further, the present invention provides a starter
composition for fermenting foods, which contains Leuconostoc
citreum CJGN 34 as an active ingredient.
[0015] In addition, the present invention provides fermented foods
such as Kimchi manufactured by using the starter compositions.
[0016] In addition, the present invention provides a lactic acid
bacteria composition containing Leuconostoc citreum CJGN 34 as an
active ingredients.
[0017] A term `active ingredient` used in this specification means
a material or materials group (including medicinal herbs containing
any unidentified pharmaceutical active ingredient) which is
contained as a main component and is expected to exhibit directily
or indirectly pharmaceutical efficacy or effect of a medicament via
inherent pharmacological action.
BRIEF DESCRIPTION OF THE DRAWINGS
[0018] FIG. 1 is a graph showing the mean of general taste scores
depending on season when Kimchi was made.
[0019] FIG. 2 is a graph showing the percentage of bacteria
isolated from fermentation bacteria flora depending on season when
Kimchi was made.
DETAILED DESCRIPTION OF THE INVENTION
[0020] The present invention is more particularly explained as
follows.
[0021] Leuconostoc citreum CJGN 34 of the present invention is
characterized by being a novel Leuconostoc citreum bacteria
isolated from Kimchi and identified. Although the bacteria was
isolated from Kimchi manufactured at Gimjang season (i.e., winter),
it does not mean that a range of the bacteria according to the
present invention is limited thereto.
[0022] According to analysis results of 16S rRNA base sequence for
identifying and classifying a microorganism, Leuconostoc citreum
CJGN 34 of the present invention has the highest homology with
Leuconostoc citreum reference bacteria (Leuconostoc citreum KCTC
3256.sup.T, GenBank accession number AF11948), which means that the
Leuconostoc citreum CJGN 34 of the present invention has the
closest molecular phylogenetic relationship with Leuconostoc
citreum. Therefore, the microorganism was identified as Leuconostoc
citreum, named as Leuconostoc citreum CJGN34 and deposited to Korea
Research Institute of Bioscience and Biotechnology (KRIBB) on Jul.
27, 2006 (Accession number: KCTC 10974BP).
[0023] Leuconostoc citreum CJGN 34 of the present invention is gram
positive bacteria and facultative anaerobe which can grow under an
aerobic or anaerobic condition. Further, Leuconostoc citreum CJGN
34 of the present invention does not form an endospore, has no
motility and has a shape of monococcus or diplococcus. Conventional
methods in the art were performed to analyze the specific shape and
physiological properties of Leuconostoc citreum CJGN 34 and the
result is summarized as following table 1.
TABLE-US-00001 TABLE 1 mannitol + sorbitol - D-mannoside -
D-glucoside + glucosamine + amygdaline + arbutin + esculine +
salicin + cellobiose + maltose + lactose - melibiose - saccharose +
trehalose + inulin - melizitose - D-raffinose - amidon - glycogen -
xylitol - gentiobiose - D-turanose + D-lyxose - D-tagatose -
D-fucose - D-arabitol - L-arabitol - gluconate - 2-gluconate -
5-gluconate - +: positive -: negative
[0024] Preferably, Leuconostoc citreum CJGN 34 of the present
invention may be stored at -70.degree. C. after being added into a
preservation liquid made of water and glycerol in an amount or be
lyophilized after being suspended in sterilized 10% skim milk, for
a prolonged stable storage.
[0025] Further, Leuconostoc citreum CJGN 34 of the present
invention, which is lactic acid bacteria isolated from
naturally-fermented Kimchi, was classified as GRAS (Generally
recognized as safe) bacteria for a human being and confirmed as
being negative in all of the safety assessment such as gelatin
liquefaction test, phenylalanine deaminase test, ammonification
test and hemolysis test, which means that the bacteria is safe as
food materials.
[0026] In case that Leuconostoc citreum CJGN 34 according to the
present invention is used in fermented foods such as Kimchi as a
starter, it may improve the general taste quality of the foods,
maintain the fermentation quality constantly throughout a year by
improving a complex sour and carbonated taste which is a
representative taste of Gimjang Kimchi traditionally manufactured
in winter and, especially, effectively inhibit deterioration of
fermentation quality in summer when taste quality is declined.
[0027] Further, a bacteria composition may be made by culturing and
harvesting bacterial flora including the bacteria. The bacteria
composition may be used according to conventional methods in
various industries such as feed probiotics after being blended with
feed materials such as grain powder; food probiotics or medicaments
probiotics in a form of tablets or capsules after being blended
with carriers or additives; or cosmetics after being blended with
the other cosmetic ingredients in an amount.
[0028] Further, Leuconostoc citreum CJGN 34 according to the
present invention may be used in a lactic acid bacteria
pharmaceutical composition. In this case, it may be formulated into
conventional pharmaceutical dosage forms known to the art. It may
be preferably formulated into oral dosage forms such as liquid,
suspension, powder, granule, tablet, capsule, bolus, pellet or
extract.
[0029] In formulating into a respective dosage form,
pharmaceutically acceptable and required carriers or additives may
be added in manufacturing the dosage form. For example, at least
one carrier selected from diluents, slip agents, binding agents,
disintegrating agents, sweetening agents, stabilizers and
preservative agents; and at least one additive selected from
flavoring agents, vitamins and antioxidants may be used in
formulating the oral dosage forms.
[0030] Any pharmaceutically acceptable carriers and additives may
be used. Specifically, it may be preferable that lactose, corn
starch, soybean oil, microcrystalline cellulose or mannitol is used
as diluents; magnesium stearate or talc is used as slip agents;
polyvinylpyrrolidone or hydroxypropylcellulose is used as binding
agents. Further, it may be preferable that carboxymethylcellulose
calcium, sodium starch glycolate, polacrilin potassium or
crospovidone is used as disintegrating agents; white sugar,
fructose, sorbitol or aspartame is used as sweeting agents; sodium
carboxymethylcellulose, .beta.-cyclodextrin, white wax or xanthan
gum is used as stabilizers; and methyl .rho.-hydroxybenzoate,
propyl .rho.-hydroxybenzoate or potassium solvate is used as
preservative agents.
[0031] Further, in addition to the above ingredients, natural herbs
with Mae-sil (japanese apricot) flavor, lemon flavor, pineapple
flavor or herb flavor, natural fruit juice, natural pigments such
as chlorophyllin or flavonoid, sweeting ingredients such as
fructose, honey, sugar alcohol or sugar, acidifiers such as citric
acid or sodium citrate may be used after mixing for a purpose of
raising appetite.
[0032] Formulating methods and carriers and additives necessary for
such formulating are detailed in Remington's Pharmaceutical
Sciences (19th ed., 1995).
[0033] Further, Leuconostoc citreum CJGN 34 according to the
present invention may be used in a lactic acid bacteria food
composition. The food composition covers conventional
daily-consumed general foods as well as health foods. If the food
composition is used in the health foods, it may be formulated into
conventional health food dosage forms known to the art, with
sitologically acceptable carriers or additives. The health foods
may be formulated, for example, into powder, granule, tablet,
capsule, suspension, emulsion, syrup, liquid, extract, jelly or
drink form. As sitologically acceptable carriers or additives, an
arbitrary carrier or additive usable in any forms to prepare may be
used.
[0034] Further, since Leuconostoc citreum CJGN 34 according to the
present invention may comprise dextran, a polymer with high
absorbability of moisture, it can be used in a lactic acid bacteria
cosmetic composition. The cosmetic composition according to the
present invention may be formulated into conventional form known to
the cosmetic industries. Any carrier or additive which is
acceptable and necessary in manufacturing a specific cosmetic form
may be added.
[0035] Further, Leuconostoc citreum CJGN 34 according to the
present invention may be used in lactic acid bacteria feed
additives or feed compositions.
[0036] Used in the feed additives, the composition may be
manufactured into a form of 20 to 90% highly concentrated liquid,
powders or granules. The feed additive may additionally include at
least one selected from organic acids such as citric acid, humalic
acid, adipic acid, lactic acid and malic acid; phosphates such as
sodium phosphate, potassium phosphate, acidic pyrophosphates and
polyphosphates (condensed phosphate); and natural antioxidants such
as polyphenols, catechins, alpha-tocopherols, rosemary extracts,
vitamin C, green tea extracts, licorice extracts, chitosan, tannic
acids and phytic acids. Used in the feed composition, the
composition may be manufactured into a conventional animal feed
form and include conventional feed ingredients.
[0037] The feed additives and the animal feed may additionally
include crops, for example, crushed or shredded wheat, oats,
barley, corn and rice; vegetable protein feeds, for example, feeds
mainly consisting of rape, soybean and sunflower; animal protein
feeds, for example, blood meal, meat meal, bone meal and fish meal;
and sugar and dairy products, for example, various dry ingredients
consisting of milk powder and whey powder, and may further include
nutritional supplements, digestion- and absorption-enhancers and
growth promoters.
[0038] The feed additives may be administered to animals
individually or in combination with other additives selected from
edible carriers. Further, the feed additives may be topdressing,
may be directly mixed with animal feeds or may be easily
administered to animals as oral dosage forms separately from animal
feeds. In case of being administered separately from animal feeds,
the feed additives may be combined with pharmaceutically acceptable
edible carriers and prepared into immediate-release formulations or
sustained-release formulations, as well known in the art. The
edible carriers may be solid or liquid, for example, corn starch,
lactose, sucrose, soy flake, peanut oil, olive oil, sesame oil and
propylene glycol. In case solid carriers are used, the feed
additives may be in a form of tablet, capsule, powder, troche or
lozenge, or may be a not-dispersed topdressing. If liquid carriers
are used, the feed additives may have a form of soft gelatin
capsules, syrup, suspension, emulsion or solution.
[0039] The feeds may include an arbitrary protein-containing
organic grain flour which has been conventionally used to meet
animals' appetite. The protein-containing grain flour typically
consists of corn or soybean flour or is a mix of corn/soybean
flour.
[0040] In addition, the feed additives and the animal feeds may
contain adjuvants such as preservatives, stabilizers, wetting
agents, emulsifiers and liquefying agent. The feed additives may be
added to animal feeds by means of dipping, spraying or mixing for
use.
[0041] The animal feeds or feed additives according to the present
invention may be applied to a diet for various animals such as
mammals, poultry and fish. The mammals may be pets (for example,
dogs, cats) as well as pigs, cows, sheep, goats and laboratory
rodents; poultry such as chickens, turkeys, ducks, geese, pheasant
and quail; and fish such trout, without limitation thereto.
[0042] As disclosed in the above, in case of novel Leuconostoc
citreum CJGN 34 according to the present invention being used as a
starter for fermented foods such as Kimchi, it may improve general
taste quality, increase a complex sour and carbonated taste which
is a unique taste of Kimchi manufactured by traditional methods in
winter and, consequently, maintain constant fermentation quality
throughout a year and effectively inhibit deterioration of
fermentation quality in summer when taste quality of Kimchi
deteriorates. Novel Leuconostoc citreum CJGN 34 according to the
present invention may be utilized as a starter for fermented foods
such as Kimchi after being prepared as a bacteria composition; as
food or medicament probiotics in a form of tablet or capsule after
being mixed with carriers or additives; or cosmetic materials after
mixing with other cosmetic materials in an amount. That is, novel
Leuconostoc citreum CJGN 34 according to the present invention may
be utilized in various industries such as medicaments, foods, feeds
and cosmetics.
[0043] Hereinafter, the present invention will be described by the
following examples in more detail. However, the purpose of these
examples is only illustrating the present invention, not limiting
the scope of the invention thereto in any way.
Example 1
Deducting Improvement Aspects by Monitoring Kimchi Quality Change
During a Year and Isolating GN34 Bacteria
[0044] Quality change of Kimchi was monitored during a year in
order to find the causes of quality change and to improve the
quality.
[0045] Kimchi manufactured according to the manufacture method of
CJ CheilJedang was used. The Kimchi manufactured between March and
May, 15 was classified as spring Kimchi; that manufactured between
May, 16 and August as summer Kimchi; that manufactured between
September and November 15 as autumn Kimchi; and that manufactured
between November 16 and February as winter Kimchi.
[0046] As a result of analyzing the taste quality of the Kimchi
manufactured in each season in a sensory test by 5-point scale, it
was found that the summer Kimchi has a low value of about 3.5 for a
general taste item, whereas the Kimchi manufactured in the other
season has an average range of 3.8 to 4.0. The above results are
shown in the FIG. 1.
[0047] In order to analyze the fermenting microbial effect on
change of the taste quality throughout a year, identification was
performed by 16S rRNA analysis according to the method disclosed in
Choi (Choi et. al., Microbial population dynamics of kimchi, a
fermented cabbage, FEMS microbial. Lett., Vol. 257, 2006, pages
262-267). The result indicates that the Kimchi manufactured during
traditional Gimjang season, i.e., in winter, has more than 80%
Leuconostoc spp. producing various acid and gas, besides lactic
acid, with sugar through a process of an absolute hetero
fermentation. Since the Kimchi manufactured in winter has the most
outstanding taste quality, it may be concluded that Leuconostoc
spp. has an important effect in providing complex fermentation
flavor and carbonated taste of winter Kimchi. On the other hand, it
was found that the ratio of Lactobacillus spp. causing partial
hetero fermentation or homo fermentation wherein only lactic acid,
not gas, is produced using sugar under a general environment, for
example, Lactobacillus sakei, Lactobacillus curvatus and
Lactobacillus plantarum was more than 50% in the Kimchi
manufactured in summer when fermentation quality is deteriorated.
The ratio is higher than that of the Kimchi manufactured in the
other seasons, which leads to a conclusion that the change of
fermentation bacteria flora may be a cause of simple sour taste and
weak carbonated taste of the summer Kimchi. The above results are
shown in FIG. 2.
[0048] In order to clarify the effect of fermentation bacteria
flora change on the fermentation flavor of Kimchi, an HPLC analysis
of organic acids was performed on the same sample as that used in
microorganism analysis, according to the methods of the Choi. In
the sample, fermentation had begun and total acidity was more than
0.4%. The organic acids correspond to main metabolites which lactic
acids produce after consuming sugar. Since lactic acid is mainly
produced in partial hetero fermentation or homo fermentation and
acetic acid and carbon dioxide as well as lactic acid are produced
as fermentation products in absolute hetero fermentation, organic
acids are major marker informing the kind of fermentation. The
analysis results are shown in Table 2.
[0049] As known from the table 2, the molar ratio of lactic acid to
acetic acid in the spring Kimchi, autumn Kimchi and winter Kimchi
was in a range of about 1.3.about.1.6, whereas that in the summer
Kimchi was in a range of about 2.2.about.2.8, which means the
proportion of Lactic acid increased. It may conclude that the
change of fermentation lactic acid bacteria increase in homo
fermentation, especially in summer, which is responsible for
quality deterioration of summer Kimchi.
TABLE-US-00002 TABLE 2 concentration ratio of Lactic manufacture
season sample No. acid/Acetic acid spring spring Kimchi 1 1.5 .+-.
0.08 spring spring Kimchi 2 1.6 .+-. 0.11 summer summer Kimchi 1
2.8 .+-. 0.13 summer summer Kimchi 2 2.2 .+-. 0.09 summer summer
Kimchi 3 2.4 .+-. 0.25 summer summer Kimchi 4 2.8 .+-. 0.13 autumn
autumn Kimchi 1 1.4 .+-. 0.09 winter winter Kimchi 1 1.4 .+-. 0.25
winter winter Kimchi 2 1.5 .+-. 0.39 winter winter Kimchi 3 1.3
.+-. 0.02 winter winter Kimchi 4 1.3 .+-. 0.09
[0050] Based on the above result, the present inventors could
manufacture Kimchi with the constant fermentation quality
throughout a year by applying lactic acid bacteria isolated from
winter Kimchi having the best taste quality, as a starter. Further,
the present inventors could isolate around 500 bacteria from winter
Kimchi in order to inhibit deterioration of the fermentation
quality of summer Kimchi. Especially, the present inventors could
isolate novel lactic acid bacteria which are absolute hetero
fermentation bacteria and dominant species of winter Kimchi and
meet the purpose of the present invention.
Example 2
Isolation and Identification of Microorganism Leuconostoc citreum
CJGN34 Bacteria
[0051] In the above Example 1, the causes responsible for the
quality change of Kimchi during a year were analyzed and lactic
acid bacteria were isolated from the well-aged delicious winter
Kimchi. The lactic acid bacteria were named as Leuconostoc citreum
CJGN34. Lactic acid bacteria Leuconostoc citreum CJGN34 were
streaked onto solid MRS medium (Difco, USA) containing 1.5% agar
and incubated at 30.degree. C. for 24 hrs. Colonies confirmed as
being purely separated were taken by a loop and incubated with MRS
broth (Difco, USA) at 30.degree. C. for 18 to 24 hrs.
[0052] Then, the morphology and physiological properties of
Leuconostoc citreum CJGN34 bacteria were determined with API50CH
and API50CHL kits (Bio-Me'reux) according to the methods disclosed
in Kim et. al., Leuconostoc inhae sp. nov., a lactic acid bacterium
isolated from kimchi, International Journal of Systematic and
Evolutional Microbiology, Volume 53, July 2003, pages 1123-1126.
The resultant morphology and physiological properties of
Leuconostoc citreum CJGN34 bacteria were summarized in the above
table 1.
[0053] Further, a sequence of 16S rRNA gene was analyzed for
identification and classification of the lactic acid bacteria. The
sequence of 16S rRNA gene was determined and analyzed according to
a method disclosed in Kim et. al., Leuconostoc kimchii sp. nov., a
new species from kimchi. International Journal of Systematic and
Evolutional Microbiology, Volume 50, September 2000, pages
1915-1919.
[0054] Since Leuconostoc citreum CJGN34 according to the present
invention has the highest homology (99.7%) with Leuconostoc citreum
reference bacteria (KCTC 3526.sup.T, GenBank accession number
AF111948), Leuconostoc citreum CJGN34 according to the present
invention was identified as Leuconostoc citreum, named as
Leuconostoc citreum CJGN34 and deposited to Korea Research
Institute of Bioscience and Biotechnology (KRIBB) Jul. 27, 2006
(Accession number: KCTC 10974BP).
Example 3
Safety Assessment of Leuconostoc citreum CJGN34
[0055] Hemolysis test, gelatin liquefaction test, hazardous
metabolite (ammonification) test and phenylalanine deaminase test
were performed according to the safety assessment methods suggested
by the standard of Korea Biotechnology Industry Organization to
assess safety of the bacteria isolated from the Example 1. The
obtained result is summarized in table 3.
TABLE-US-00003 TABLE 3 Safety assessment for Leuconostoc citreum
CJGN34 tests gelatin phenylalanine bacteria liquefaction deaminase
hemolysis ammonification CJGN34 negative negative negative
negative
[0056] Based on the above result, Leuconostoc citreum CJGN34 was
found negative in all of the gelatin liquefaction test, hazardoug
metabolite (ammonification) test, phenylalanine deaminase test and
hemolysis test. Accordingly, Leuconostoc citreum CJGN34 was
confirmed as being safe for administration to a human being.
Example 4
Characteristics of Kimchi Manufactured by Using Leuconostoc citreum
CJGN34 as a Starter
[0057] A bacteria culture or a bacteria composition was prepared as
follows to apply Leuconostoc citreum CJGN34 according to the
present invention as a starter.
[0058] Culture medium was prepared by dissolving 10 g of glucose, 5
g of yeast extracts and 5 g of diammonium citrate into water,
sterilizing, inoculating with bacteria and incubating at
25.about.37.degree. C. for 8 to 24 hrs. A bacteria composition was
prepared as follows: bacteria were incubated and grown at
30.degree. C. for around 18 hrs with pH adjustment of MRS broth
(Difco) into 5.5.about.7.0 by ammonia gas and centrifuged for
recovery. Recovered bacteria were lyophilized below -40.degree. C.
with a cryoprotectant consisting of dextrin 5% and skim milk 10%,
dried at the temperature of 30.about.37.degree. C. and ground into
powder by a mixer. Powdered viable cells were counted to a desired
value, mixed with a suitable amount of excipients such as glucose,
lactose and skim milk and packaged in a sealable aluminium
pouch.
[0059] The culture medium or the bacteria composition manufactured
according to the above was added in an amount of 0.001%.about.3%
based on the total weight of Kimchi in manufacturing Kimchi. Kimchi
was fermented and aged according to the manufacturing method of CJ
Cheiljedang. Further, a sensory test and physiochemical analyses
were performed with two (2) kinds of Kimchi made of the same raw
materials and composition ratio, one of which was fermented with a
starter Leuconostoc citreum CJGN34 and the other of which was
fermented naturally without a starter. Since the result may be
different depending on aging, the aging was adjusted within a range
of 0 to 0.05% based on the total acidity. The sensory test result
for the Kimchi manufactured in the above is shown in table 4.
TABLE-US-00004 TABLE 4 General carbonated manufacture taste
Sourness taste Aftertaste season sample preference preference
preference preference Summer naturally 3.59 3.22 3.49 3.36
fermented Kimchi Summer starter 3.85 3.67 3.61 3.81 Kimchi Winter
naturally 4.01 3.81 3.71 3.76 fermented Kimchi Winter starter 4.05
3.92 3.82 3.79 Kimchi Spring naturally 3.81 3.66 3.68 3.68
fermented Kimchi Spring starter 3.91 3.71 3.75 3.72 Kimchi Autumn
naturally 3.86 3.72 3.66 3.71 fermented Kimchi Autumn starter 3.89
3.78 3.71 3.7 Kimchi
[0060] As shown from the table 4, general taste preference for the
naturally fermented Kimchi shows a wide deviation depending on
manufacture season, i.e., a value of 3.59 for the summer Kimchi to
a value of 4.01 for the winter Kimchi. On the other hand, the
Kimchi manufactured with a starter Leuconostoc citreum CJGN34 shows
a narrower deviation of the taste quality, i.e., a value of 3.85
for the summer Kimchi to a value of 4.05 for the winter Kimchi,
which means that Leuconostoc citreum CJGN34 has a significant
effect in providing Kimchi with a constant fermentation quality
throughout a year. Especially, the naturally-fermented summer
Kimchi shows lower values for sourness preference, carbonated taste
preference and aftertaste preference with a lower value of about
3.5 for general taste preference. However, the starter Kimchi has a
variation of sourness preference and aftertaste preference within
95% significance, a variation of carbonated taste preference within
90% significance and a variation of general taste preference within
95% significance, which means that deterioration of fermentation
quality of the summer Kimchi was effectively inhibited.
[0061] The molar ratio of lactic acid to acetic acid which is a
main remark showing fermentation type among hetero fermentation and
homo fermentation shows that the starter summer Kimchi has a lower
value compared to the naturally-fermented summer Kimchi. Such
result indicates that the starter Leuconostoc citreum CJGN34
normally dominated, inhibited homo fermentation- or hetero
fermentation-lactic acid bacteria and effectively controled
deterioration of Kimchi fermentation quality.
[0062] The result analyzing organic acids of the Kimchi
manuracfured in each season is shown in a following table 5.
TABLE-US-00005 TABLE 5 manufacture concentration ratio of lactic
season sample acid/acetic acid (mM) summer naturally fermentation
Kimchi 2.4 .+-. 0.3 summer starter Kimchi 1.5 .+-. 0.1 winter
naturally fermentation Kimchi 1.4 .+-. 0.2 winter starter Kimchi
1.3 .+-. 0.3 spring naturally fermentation Kimchi 1.5 .+-. 0.1
spring starter Kimchi 1.5 .+-. 0.1 autumn naturally fermentation
Kimchi 1.4 .+-. 0.1 autumn starter Kimchi 1.4 .+-. 0.1
Example 5
Fermented Foods and Pickles Manufactured Using Leuconostoc citreum
CJGN34 Bacteria as a Starter
[0063] For applying Leuconostoc citreum CJGN34 bacteria according
to the present invention to fermentation foods besides Kimchi as a
starter, a bacteria culture or a bacteria composition was prepared
in the same manner with the Example 4 and used as a starter in
manufacturing fermented foods. Fermented foods were made of various
raw materials such as cucumber, radish, carrot, onion, garlic,
pepper and Korean cabbage. Further, fermented picked radishes were
manufactured by applying the starter to picked radishes.
Example 6
Preparation and Usage of a Bacteria Composition Containing
Leuconostoc citreum CJGN34
[0064] Leuconostoc citreum CJGN34 identified in the Example 1 was
mass produced and a bacteria composition was prepared according to
the Example 4 as materials of medications, foods, feeds, feed
additives or cosmetics.
[0065] The prepared bacteria composition was used as feed
probiotics after being mixed with feed materials such as grain
powder; as medications or food probiotics in a form of tablets or
capsules after being mixed with carriers or additives; or as
cosmetic materials after being mixed with the other cosmetic
materials. The bacteria composition could be utilized in various
industries according to conventional methods in the art.
* * * * *