Trife Inc. Oralpeace Project
Bashamichiotsu Building 4-43 Minaminakadōri, Naka-ku, Yokohama
PRESS RELEASE（2017/7/28）

The evolution of oral care agents that are safe to swallow Antibacterial effect against oral candida of the natural antibacterial agent
derived from vegetable lactic acid bacteria peptides (protein) androse essential oil- Covers bacteria causing infectious diseases in the oral cavity[Overview]
Trife Inc. (Headquarters: Yokohama, Yokohama Business Grand Prix 2014 BestCompany), through industrial-academic collaboration with Kyushu University Facultyof Agriculture (Fukuoka Prefecture) and Yasashii Laboratory Inc. (FukuokaPrefecture), has confirmed that the natural antibacterial peptide (protein) Nisin A (*1),
produced by vegetable lactic acid bacteria, in an original combination of Damask Roseessential oil from Bulgaria to form the natural antibacterial agent Neonisin-e, hasstrong antibacterial effect against the fungus (yeast) which causes oral candidiasis(*2).
This research group announced Neonisin at the Kyushu University Press Club in
December 2012, as a natural antibacterial agent which combines high-purity Nisin Awith plum extract in a unique ratio. It is a natural formulation that is made of only edible
ingredients and is safe to swallow, which has gained attention as the world's first
formulation with excellent bactericidal effect on gram-positive and gram-negative
bacteria which are causative of dental caries, periodontal disease, and aspiration
pneumonia. However, it had a weak point in that it was not effective against fungi(yeast), which is a causal organism for oral candidiasis.
Now Neonisin-e at this time expands the antibacterial spectrum to not only thebacteria of the conventional Neonisin, but also to fungus (yeast), and this lactic acidbacteria peptide preparation which is safe to swallow will cover most of the pathogenic
bacteria that cause infections in the oral cavity. In the autumn of this year, we areplanning to release products covering the causal bacteria responsible for infections in
the oral cavity, effective not only for dental caries, periodontal disease and aspiration
pneumonia, but also for oral candidiasis, which is commonly found in elderly people,infants, disabled people and cancer patients with weak immunity.
[Background]
Both dental caries and periodontal disease are infections caused by bacteria, it issaid that 90% of Japanese people have dental caries and 70% suffer from periodontal
disease. In addition, increase in bacteria in the oral cavity is major problem as a risk
factor for aspiration pneumonia (*3), which is the cause of death of as many as 300
people a day, mainly elderly people.
Candida is a fungus that lives on human skin and mucous membranes, and cancause infections in people with an underlying disease (cancer, blood disease,immunodeficiency disease such as AIDS, diabetes etc.), or who are taking

immunosuppressive agents or antibiotics, and in infants and elderly people who haveweak immunity. Especially in the elderly, oral candidiasis (*2), in which the surface ofthe tongue is covered with white spots, is often a problem.
Treatment of oral candidiasis is usually done by oral care, and the use of antifungal
agents (potent synthetic fungicides) or natural formulations such as tree-derived
essential oils which cause concerns about safety if swallowed. However, oral carealone cannot disinfect or cure oral candidiasis once it has developed, and since theuse of antifungal drugs and conventional natural preparations has problems withresistant bacteria and side effects (such as stomach trouble, teratogenic effects), newtreatments that are effective and safe even when swallowed have been sought all overthe world.
[Content and Research Results]
1 The Development and Practical Application of Neonisin
Discovered by Professor Kenji Sonomoto of Kyushu University Faculty ofAgriculture, Department of Bioscience and Biotechnology, the leading researcher on
lactic acid bacteria, Nisin A, a natural antibacterial peptide (protein) produced by lactic
acid bacteria in okara, and a technology to extract it at high purity was developed by abio-venture over 10 years of research. Furthermore, in 2012 we developed a lactic acidbacteria peptide preparation "Neonisin” (Patent No. 5750552) that combines our own
high purity Nisin A with plum extract, which exhibits excellent bactericidal effect against
oral disease bacteria that cause dental caries (Streptococcus mutans, Gram positivebacteria) (*4) and periodontal disease-causing bacteria (Aggregatibacter
actinomycetemcomitans, Gram negative bacteria) (*5). In July 2013, it was launchedas the oral care product Oralpeace, which contains Neonisin, and the world's firstpractical application of Neonisin was achieved.
2. Development of Neonisin-e
Neonisin shows excellent antibacterial effect, but had a weak point in that it is lesseffective against fungi (yeast). Therefore, in order to compensate for the weak point ofNeonisin, selection tests of various natural substances were conducted, and as aresult, we found a synergistic effect with Nisin against yeast in a small amount ofDamask rose essential oil. Although it was said that Damask rose essential oil has
antibacterial activity, a certain concentration is necessary, and little effect was seen intrace amounts. However, the lactic acid bacteria peptide preparation Neonisin-e, which
combines high-purity Nisin with a small amount of Damask rose essential oil in aunique formulation ratio, showed enhanced antibacterial effect. Neonisin-e is anevolved version of the previous Neonisin. This lactic acid bacteria peptide formulation
which is safe to swallow will cover most bacteria that cause infectious diseases in theoral cavity.
[Features of Neonisin-e]
1. Excellent antibacterial activity against candida, dental caries and periodontal
disease bacteria
2. Highly safe with natural origin
3. Excellent biodegradability and safety after decomposition (environmentally andhuman friendly)Synergistic Effect of Antibacterial Activity(Antibacterial peptide + rose oil)

Antibacterial Activity (Candida)
Antibacterial peptide
Rose oil

Antibacterial peptide+rose oil

Figure 1. Antibacterial activity of antibacterial peptide (Nisin A) + roseoil (Damask rose essential oil) against yeast

Figure 2. Antibacterial activity of the lactic acidbacteria peptide formulation Neonisin-e against yeast

Table 1. Antibacterial Activity of the Lactic Acid Bacteria Peptide Formulation
Neonisin-e (Rose Oil) against Bacteria and Fungi
Antibacterial Activity Test (1-minute sterilization rate)

Without

Rose oil

57100

10100

55

9956100

985

Candida yeast

17100

Candida yeast

45100

Gram negative
periodontal disease
Gram negative
periodontal disease
Gram positive
dental cariesGram positive
dental caries
Candida yeast(Data provided by Kagoshima University Graduate School of
Medical and Dental Sciences, Komatsuzawa Laboratory)[Future Development]
Neonisin-e is a new 100% natural antibacterial agent that combines high-purity
antibacterial peptides (Nisin A) produced by the lactic acid bacteria with Damask rose
essential oil in a unique ratio. While safe to swallow, it is expected to be usedeffectively as a natural ingredient with antibacterial effects on dental caries, periodontal
disease bacteria and candida. This autumn, we plan to launch an oral care productcontaining Neonisin-e for people suffering from infections in the oral cavity.

Reference Materials
Table 2. Comparison of Oral Antibacterial Agents

Dental caries /Aspiration pneumonia

Gram negativebacteria
Periodontaldisease

Fungus (yeast)
CandidaNaturalsubstance

Alcoholr´Cetylpyridinium
chloride (CPC)r´

Chlorhexidine (CHX)´´Benzethonium chlorider´

Hypochlorous acidwater´Lactic acid bacteriarr´

Plant extractrr´

Hinokitiol (*)

○○

Nisin A´´Neonisin´Neonisin-eAntibacterialcomponent

Gram positivebacteria(*) Strong odor affects taste ´

Suspected teratogenicity ´[Glossary]*1 Nisin ANisin A is an antibacterial peptide (protein) produced by lactic acid bacteria(Lactococcus lactis), consisting of 34 amino acids. Nisin is naturally contained in
traditional fermented foods such as yoghurt, cheese and rice bran pickles, whichpeople have eaten since ancient times. It was discovered in 1928, and since it wasapproved by WHO and FAO in 1969, it has been used in food as a safe preservative inover 50 countries around the world. In Japan, it was approved as a food additive by theMinistry of Health, Labour and Welfare in 2009. Nisin A is water soluble and exerts its
effect at a concentration of one part per billion. All negative results have been obtainedin genotoxicity, carcinogenicity and other tests, and its safety has been demonstrated.
It is easily broken down and digested with enzymes in the human intestinal tract, and isbiodegradable so that release to the natural world is possible without environmentalburden.

Structural formula of Nisin A

3D molecular model of Nisin A

*2 Oral Candidiasis
Oral candidiasis is an opportunistic infection caused
by Candida, a resident fungus in the oral cavity. It iscaused by systemic factors such as an underlying
disease (such as cancer, a blood disease,immunodeficiency disease such as AIDS, or diabetes),
the weak immunity of infants and elderly people, ortaking immunosuppressive agents or antibiotics, andappears as milky white mossy spots in mucous(Photo: Wikipedia)
membranes. Treatment of oral candidiasis is usually
done by oral care, and the use of antifungal agents (potent synthetic fungicides) or
natural formulations such as tree-derived essential oils which cause concerns about
safety if swallowed. However, oral care alone cannot disinfect or cure oral candidiasis
once it has developed, and since the use of antifungal drugs and conventional naturalpreparations has problems with resistant bacteria and side effects (such as stomachtrouble, teratogenic effects), new treatments that are effective and safe even whenswallowed have been sought all over the world.
*3 Aspiration Pneumonia
Aspiration pneumonia is pneumonia caused by bacteria flowing into the lungs withsaliva and gastric juice. It occurs frequently in the elderly, and is characterized byrepeated recurrence. As recurrence is repeated, bacteria develop resistance toantibiotic treatment, leading to the deaths of many elderly people. Currently,pneumonia is the third leading cause of death in Japanese. About 95% of those who
die are over 65 years old, and prevention is being called for.
*4 Streptococcus mutans
Streptococcus mutans is a kind of gram-positive streptococci that causes dentalcaries. This bacterium breaks down sucrose and creates a sticky substance called
insoluble glucan (biofilm). It proliferates in the biofilm, produces acid, and causes
dental caries.
*5 Aggregatibacter actinomycetemcomitans
Aggregatibacter actinomycetemcomitans is a kind of gram-negative bacteria that isa cause of periodontal disease. It also has virulence that affects tissues such as whiteblood cells.
*6 Candida albicans
Candida albicans is a kind of yeast that is a cause of oral candidiasis. Originallyliving in the human body surface, gastrointestinal tract and vaginal mucosa, it canbecome an opportunistic infection which causes lesions when the physical condition
becomes poor.

The Oralpeace Project
The research and development of Oralpeace was based in the world’s leading lactic
acid bacteria biotechnology research of government, industry and academia.
Released in July 2013, Oralpeace is an oral care product containing the globally
innovative patented formulation Neonisin, that is safe to swallow while having excellent
disinfection effect on trouble-causing bacteria in the oral cavity. For reducing nursing
care expenses of the elderly, which exceed 9 trillion yen in Japan, and efforts to create
work and income for disabled people nationwide, it was awarded the Yokohama
Business Grand Prix 2014 Grand Prize and the highest Japan Venture Award 2015. It
is attracting attention as a bio-social venture that is leading Japan and Yokohama indriving the world's aging society, welfare and health care industry. The project featuresa structure of three employees and the efforts of more than 100 highly motivated pro
bono members.

[Contact]
Trife Inc.
Bashamichiotsu Building 4-43 Minaminakadōri, Naka-ku, Yokohama, Japan 231-0006
In charge: Tejima/Ueda TEL: 045-663-2101
Mail: contact@oralpeace.com
Website: http://oralpeace.com/ Facebook: https://www.facebook.com/oralpeace