Table Of ContentANTIBIOTIC SUBSTANCES PROM YEASTS
A Thesis
Presented to
the Faculty of the Department of B acteriology
U niversity of Southern C alifo rn ia
In P a rtia l F u lfillm en t
of the Requirement fo r the Degree of
M aster of Science
by
M itsuru Nakamura
June 1950
UMI Number: EP55015
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This thesis, written by
MITSURU ..NAKAMURA
under the guidance of h%3.... Faculty Committee,
and approved by all its members, has been
presented to and accepted by the Council on
Graduate Study and Research in partial fulfill
ment of the requirements for the degree of
MASTER OF SCIENCE
Date 1M M ..1950
Faculty Committee
TABLE OF CONTENTS
CHAPTER PAGE
I . THE PROBLEM ......................................... 1
Statem ent of the p r o b le m ............................... 1
Importance of the s t u d y .................................... 1
I I . REVIEW OF THE LITERATURE.......................... 3
I I I . METHODS AND RESULTS............................... 15
EXPERIMENTS................................................ 15
Screening of y easts fo r a n tib io tic a c ti
v ity by ad ju stin g thep H ............................. 16
Best carbon source fo r a n tib io tic pro
duction by Torula u t il i s . . . . . . 19
Best nitrogen source fo r a n tib io tic
production by T. u t i l i s .............................. 20
E ffect of shaking the cu ltu re on a n tib i
o tic production .................................... 22
E x tractio n of the a n tib io tic substance
from the m edium ........................................ 23
Use of sy n th etic media . . . . . . . . 26
O btaining a b a c te ria l spectrum fo r the
a n tib io tic . . . . . 31
T esting fo r heat s ta b ility of the
a n tib io tic . . . . . 31
Toxicity te s ts on r a t s .................................... 34
H i
CHAPTER PAGE
XV. SUMMARY AND CONCLUSIONS' . . . . . . . . . . . . 35
BIBLIOGRAPHY ............................................................................................. 38
LIST OP TABLES
TABLE PAGE
I. The Composition of U niversal y east Medium*. . 15
II. Yeasts Screened fo r A n tib io tic A ctiv ity . . . 15
I I I . A n tib io tic A ctiv ity of F iltra te s of Y easts
Grown a t Varying PH Levels A gainst S. aureus 17
IV. A n tib io tic A ctiv ity of F iltra te s of Y easts
Grown a t Varying PH Levels A gainst E. o o li 17
V. Further Studies on the E ffect of PH on the
Production of A n tib a c te ria l Substances . . 18
VI. E ffect of Carbon Source on A n tib io tic
Production by Torula u t il i s . . . . . . . 20
V II. E ffect of N itrogen Source on A n tib io tic
Production by Torula u t i l i s . ..................... 22
V III. Measurement of In h ib itio n Produced by E ther
E x tract of T. u t il i s medium . . . . . . . 24
IX. Compositions of S ynthetic Media Used fo r
A n tib io tic Production • • • • • • • • • • 27
X. End-Products of Yeast Ferm entation of Sugar in
R elative Proportions U sually Present . . . 50
XI. P ro p erties of E n d -P ro d u c ts ...................................... . 50
X II. B a cterial Spectrum Obtained from an A n tib io tic
Produced by T. u t il i s on S ynthetic Medium 52
CHAPTER I
THE PROBLEM
A n tib io tics are playing an im portant ro le in the fie ld
of m edical sciences today. Many a n tib io tic s have been is o
la te d and described from b a c te ria , molds, and streptom yces,
but very l i t t l e work has been reported on the p o s s ib ility of
producing an a n tib io tic substance from y e a sts.
Statem ent of the problem , The purpose of th is study
was to survey ten yeasts fo r the production of an a n tib io tic
p rin c ip le . If a y east was found th a t formed an a n tib a c te ria l
agent, the aim was to determ ine the optimum conditions fo r
maximum a n tib io tic production, and to is o la te the agent so
th a t d e ta ile d stu d ies of i t s spectrum could be obtained.
Importance of the stu d y . D espite the fa c t th a t many
a n tib io tic s have been discovered, re la tiv e ly few are used in
m edicine. Unless the newly discovered agent dem onstrates a
m arkedly d iffe re n t spectrum again st pathogens i t is not
fe a sib le to develop i t com m ercially in com petition w ith
compounds fo r which the fac to rs of production are known.
The fa c to rs c o n tro llin g the production of y easts are
more fu lly understood than fo r any o th er group of organism s.
A good a n tib io tic from th is source could be produced cheaply
and in large q u a n titie s. I t was hoped th a t in searching fo r
2
a new a n tib io tic fronr y east, one m ight be* found th a t would
be e ffe c tiv e ag ain st a wide v a rie ty of organism s.
CHAPTER II
REVIEW OP THE LITERATURE
For many years there has heen a popular theory th a t
abscesses and pimples w ill disappear when y east is ingested
and th a t i t was of im portance also in the treatm ent of other
skin d iseases. Long before s c ie n tis ts re a liz e d th a t y east
contained vitam ins, physicians were using i t fo r therapy
ag ain st skin d iso rd e rs.
The a n tib a c te ria l a c tiv ity of the y east was f i r s t
rep o rted by Sergent (1903), who rep o rted a substance p ro
duced by yeast which was toxic to Staphylococcus.aureu s.
B urnett (1905) reported the use of yeast as a d isin fe c ta n t.
Hayduck (1909) m entions in h is paper th a t Lange, Henneberg,
and Hayduck have d etected the presence in c e rta in cereals
of substances th a t are poisonous to yeast when dissolved
to g eth er w ith sugar in d is tille d w ater. He was able to
o b tain a sim ila r substance from d i s t i l l e r ’s y east when i t
was dried rap id ly a t high tem peratures, and ex tracted w ith
w ater containing 0.1 per cent hydrochloric acid . He found
th a t no such poisonous e x tra c t is obtainable from yeast
th a t has been dried slow ly, owing probably to the actio n of
try p tic enzymes upon the poisonous p ro te in . He ex tracted a t
both 25° C. and a t 50° G., but found th a t an e x tra c tio n a t
50° C. gave b e tte r re s u lts . At 50° C., the e x tra c tio n is
4
completed in two hours. His experim ents in d icated th a t n e ith e r
su lfu ric acid nor a c e tic acid can replace hydrochloric acid
in th is e x tra c tio n process. To prepare a poisonous e x tra c t
from brewer*s y east, he found th a t i t was necessary to remove
some of i ts lime by d ilu te acid previous to the e x tra c tio n
proper. Calcium s a lts prevent the poisonous e ffe c t. If the
y east was weakened by storage before being d ried , a le ss
poisonous e x tra c t re su lte d . The a ctio n of th is so lu tio n was
in te n si fied by d ia ly s is .
L ater in the same year, Fernbaeh (1909), using approxi
m ately the same procedure as Hayduck (1909), found an activ e
substance th a t could k i ll b a c te ria . He ex tracted a m acera
tio n of d essicated y east w ith d ilu te (1:1000) hydrochloric
acid fo r 19 hours a t 37° C ., and f ilte r e d the so lu tio n . A fter
n e u tra liz a tio n , the f i l t r a t e exh ib ited marked to x ic ity to the
yeast i ts e l f as w ell as to various o th er stra in s and to
b a c te ria . In h is experim ents he used one Bacterium c o ll and
one Staphylococcus aureus pyogenes s tra in . The so lu tio n
proved to be b a c te ric id a l to both of these, though somewhat
more inten se ag ain st the la tte r . He found the toxin to be
v o la tile . Upon d is tillin g the f i l t r a t e under dim inished
pressure such th a t the tem perature did not ris e above 40° C.,
the to x ic ity was tra n sfe rre d to the d is tilla te w hile the
re sid u a l liq u id was in a c tiv e . No reactio n s were obtained fo r
aldehydes—notably form aldehyde--to which b a c te ric id a l actio n
