# US 4,396,763 A · High molecular polysaccharide MPS-80

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- Number: US 4,396,763 A
- Document ID: US4396763A
- Status: Granted
- Kind: A
- Application number: 06/322,183
- Filed: 1981-11-17
- Granted: 1983-08-02
- Earliest claimed: 1981-01-14
- Assignees: Meiji Milk Products Co., Ltd.
- Inventors: Fumiyasu Tsuchiya; Kyushichi Miyazawa; Michio Kanbe; Munehiro Oda; Naoyuki Ebisawa
- Web page: https://patentgrep.com/patent/US4396763A

## CPC

- A61K 31/715: Polysaccharides, i.e. having more than five saccharide radicals attached to each other by glycosidic linkages; Derivatives thereof, e.g. ethers, esters
- C12P 19/04: Polysaccharides, i.e. compounds containing more than five saccharide radicals attached to each other by glycosidic bonds
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)}

## Abstract

New high molecular polysaccharide MPS-80 was obtained by culturing high molecular polysaccharide MPS-80-producing bacteria belonging to genera Lactobacillus or Streptococcus. The new high molecular polysaccharide MPS-80 has an excellent anti-tumor effect, and it is expected for the utilization as anti-tumor agent.

## Claims

### Claim 1

High molecular polysaccharide MPS-80 having the following physicochemical properties:
(1) elementary analysis:
C: 42.2%
H: 6.9%
O: 50.4%
(2) molecular weight:
(i) molecular weight determined by ultrafiltration method by means of Sepharose 2B as shown in FIG. 1;
said polysaccharide is fractionated near the void volume under conditions comprising 2.5.times.40.5 cm column size, 5 g fraction, 2.5 mg (1 ml) sample loaded and 0.05 M phosphate buffer solutin (pH 6.0) used as developer;
(ii) molecular weight determined by ultracentrifugation wherein
a sample is dissolved in 0.2 M phosphate buffer solution (pH 7.3) and the resulting solution having a concentration of 0.1% is subjected to the ultracentrifugation (preset rotation rate: 51,200 RPM) to obtain a sedimentation constant of 7.98S (S: Svedberg unit), the sedimentation pattern being single;
(3) melting point (decomposition point):
said polysaccharide begins to discolor at around 262.degree. C. and is blackened at 263.degree.-264.degree. C.
(4) specific rotatory power:
[.alpha]sub.D.sup.18 =+33.2 (C.dbd.0.5%)
(5) ultraviolet ray absorption spectrum:
as shown in FIG. 2.
(6) infrared ray absorption spectrum:
as shown in FIG. 3.
(7) solubilities in solvents:
soluble in water but insoluble in methanol, ethanol, acetone and ether;
(8) color reactions:
(i) Molisch reaction: +
(ii) Anthrone reaction: +
(iii) Cysteine sulfuric acid reaction: +
(iv) Aniline hydrochloric acid reaction: -
(v) Carbazole sulfuric acid reaction: -
(vi) Elson-Morgan reaction: -
(vii) Biuret reaction: -
(9) basicity, acidity or neutrality:
0. 1-0.5% aqueous solution of said polysaccharide has a neutral pH;
(10) color:
freeze-dried polysaccharide is in the form of white fibers;
(11) saccharide constituting the polysaccharide are glucose and galactose determined by examining said polysaccharide by GLC using 5% SE-52 (2 m column) as follows:
the sample was hydrolyzed with 2 N-H.sub.2 SO.sub.4 in boiling water for 4 hours, then neutralized with barium carbonate and subjected to the filtration; the filtrate was demineralized with Amberlite IRA-410 and Amberlite IR-120B, concentrated to dryness and subjected to TMS treatment followed by GLC;
(12) proportion of saccharide constituting the polysaccharide:
a sample of said polysaccharide was hydrolyzed with 2N-H.sub.2 SO.sub.4 in boiling water for 4 hours, neutralized with barium carbonate and subjected to filtration; the filtrate was examined to determine a proportion of saccharide constituting the polysaccharide by enzyme method; the ratio of glucose to galactose was 2.2-1.9 to 1;
(13) C.sup.13 -NMR Spectrum (in D.sub.2 O, TMS standard) (ppm):
as shown in FIG. 4;
(14) decomposition with enzymes:
said polysaccharide was dissolved in 0.05 M acetate buffer solution and subjected to reaction with various enzymes; the decomposition by the enzymes was judged from increase in amount of reduced sugar by Somogyi-Nelson method; Enzymes used and reaction conditions:
______________________________________ a. Amylase (a product of Boehringer) pH 5.9, 37.degree. C., 4 hrs. b. Amylase (a product of Boehringer) pH 4.8, 30.degree. C., 4 hrs. c. Galactosidase (a product of Boehringer) pH 4.8, 30.degree. C., 4 hrs. d. Amyloglucosidase (a product of Boehringer) pH 4.8, 30.degree. C., 4 hrs. e. Galactosidase (a product of Boehringer) pH 4.8, 30.degree. C., 4 hrs. ______________________________________
no increase in amount of reduced sugar was recognized at all under above conditions a through e;
(15) LD.sub.50
ddY w.female. mice (average body weight: 21.3 g; each group consisted of 7 mice) were used; a sample solution in physiological saline was administered intraperitoneally once in various amounts to the mice and they were observed for 10 days to determine LD.sub.50 ;
as a result, LD.sub.50 was not less than 200 mg/Kg-body weight; and
(16) MPS-80 is produced by cultivating a member selected from the group consisting of Lactobacillus Jugurti No. 851, FERM BP-66 and Streptococcus thermophilus No. 127, FERM BP-65.