Attached
3 strains of fungi & 4 strains of bacteria that showed
cellulolytic activity were isolated froftlsoil & Ii strains of
ba.cteria were isolated aerobically fran cowdung. 18 strains
of bacteria were isolated anaerobically from the rumencontents of
cattle. Majorpart of this investigation was carried
out with anaerobes isolated fran the rumencontents.
The cellulosic substrate used wasmicrocrystalline
cellulose (Bee), which is the crystalline portion of
cellulose. Dueto its insoluble nature bowever, carboxymethylcellulose (CMC)had to be substituted as the sole
carbon source for sane of the growthexperiments. The
applicability of most of the growth studies wastested
usinq -a strain of Ruminococcusalbus which is knosn to
require strict anaerobic conditions for growthand is
excellent in cellulose digestibility & the production of
ethanol.
The cellulolytic ability of the strains isolated
fran roil was tested by visual observation of cellulose
degradation that included filter paper decay & the formation
of cleared zones in agar medium& by the measuranentof
total carbohydrate content of t.hemediumby the phenolsulphuric acid colourimetric test. The irolates fran
cowdungwere tested by the phenol-sulphuric acid test
& chanical analysis of the culture broth. The strains
isolated fran the rumencontents were tested by the phenol
sulphuric acid test.
Althoughisolated fran cultures that showed
cellulolytic actvity only two soil isolates showed
clearing on agar medium. Four strains showedsignificant
cellulolytic activity by the phenol-sulphuric acid test. Three irolates fran cow dung showed filter paper activity
while gas production was observed by only one isola te. Of
the isolates fran the rumen contents, only nine strains
showedsignificant cellulolytic activity by the phenol -
sulphuric acid test.
For analytical studies, the anaerobic condition
of the mediumwas obtained by culturing yeast in the medium,
prior to inoculation of the anaerobes. Experiments carried
out using R. albus, & twJ isolates indicated that the
presence of yeast in the rned.i.urndid not affect
growth & cellulose digestibility of the anaerobes.
Chemical analysis of the distillate of the culture
broth of R. albus indicated positive results for ethanol
& sugars. Quantitative analysis of ethanol was done by gas
liquid chrcmatcqraphy. Sugars were separated by paper
chromatography with isoproPanol-pyridine-acetic acid -
water & glucose was quan~itatively analysed by the phenolsulphuric acid test.
9.2 mg. ethanol per 100 ml. broth was produced by
R. albus in 1% 0'1: mediumat 10 day incubation at 37 'C.
The production of sugars increased upto a maximumof 30 mg.
per 10) ml. broth at 6 days & decreased thereafter. Cell
growth assessed by optical density measurements of the
culture showed a maximumat 6 days. The maximumarnounts
of ethanol & sugars produced fran 1% cellulose broth were
1.75 mg. & 15.48 mg. per 100 mi. broth respectively.
The effect of sane envirorrnental factors on the
product.Lon of ethanol was studied using cultures of R.albus.
Of these factors, rraintenance of pH of the broth at 6.8,
incubation under CO2
gas & seed cultivation of bacteria in
0.03% cellobiose broth favoured production, while incubation
at room temperature with shaking reduced production.A study of growth curves of the isolates sbowed
a harvest time of 10 days as suitable for obtaining samples
for analysis for ethanol. The pattern of pH variation of
each culture studied with the pattern of cell growth, indicates
the enzymic activity of the relevant isolate.
The negligible arrounts of ethanol produced by the
isolates from cellulose have increased under favourable
circumstances. Four isolates yielded ethanol fran eM::::
while three others have yielded ethanol from cellulose.
Glucose was detected in the culture broth of tv.o isolates
only. Cello1!liose was detected in the cultures of four isolates.
An unidentified trimer was producer by all isolates except
one.
The production of ethanol was increased by mixed
cultures, carried out using the canbinations of ethanol
producers with non-ethanol producers. A further increase
in the production was observed using combinations of ethanol
producers, coupled with the Cl & ex components. 56 mg. of
ethanol Per 100 mI. broth, the highest arrount obtained
in this study was produced by the combination of R. albus
& an isolate. Al.t.bouqhrrost of the canbinations favoured
production one combination did not produce ethanol.
However, the cell yields & the production of
ethanol & sugars obtained in this study are fairly low1
partly due to the nature of the cellulosic substrate used.
Hence, the resul ts obtained are rrore of an acadanic interest
rather than of econanic value.
