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Abstract:
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The present study aimed at the utlisation of microbial organisms for the
production of good quality chitin and chitosan. The three strains used for the
study were Lactobacillus plantarum, Lactobacililus brevis and Bacillus subtilis.
These strains were selected on the basis of their acid producing ability to reduce
the pH of the fermenting substrates to prevent spoilage and thus caused
demineralisation of the shell. Besides, the proteolytic enzymes in these strains
acted on proteinaceous covering of shrimp and thus caused deprotenisation of
shrimp shell waste. Thus the two processes involved in chitin production can be
affected to certain extent using bacterial fermentation of shrimp shell.Optimization parameters like fermentation period, quantity of inoculum,
type of sugar, concentration of sugar etc. for fermentation with three different
strains were studied. For these, parameters like pH, Total titrable acidity (TTA),
changes in sugar concentration, changes in microbial count, sensory changes
etc. were studied.Fermentation study with Lactobacillus plantarum was continued with 20%
w/v jaggery broth for 15 days. The inoculum prepared yislded a cell
concentration of approximately 108 CFU/ml. In the present study, lactic acid and
dilute hydrochloric acid were used for initial pH adjustment because; without
adjusting the initial pH, it took more than 5 hours for the lactic acid bacteria to
convert glucose to lactic acid and during this delay spoilage occurred due to
putrefying enzymes active at neutral or higher pH. During the fermentation study,
pH first decreased in correspondence with increase in TTA values. This showed
a clear indication of acid production by the strain. This trend continued till their
proteolytic activity showed an increasing trend. When the available sugar source
started depleting, proteolytic activity also decreased and pH increased. This was
clearly reflected in the sensory evaluation results. Lactic acid treated samples
showed greater extent of demineralization and deprotenisation at the end of
fermentation study than hydrochloric acid treated samples. It can be due to the
effect of strong hydrochloric acid on the initial microbial count, which directly
affects the fermentation process. At the end of fermentation, about 76.5% of ash was removed in lactic acid treated samples and 71.8% in hydrochloric acid
treated samples; 72.8% of proteins in lactic acid treated samples and 70.6% in
hydrochloric acid treated samples.The residual protein and ash in the fermented residue were reduced to
permissible limit by treatment with 0.8N HCI and 1M NaOH. Characteristics of
chitin like chitin content, ash content, protein content, % of N- acetylation etc.
were studied. Quality characteristics like viscosity, degree of deacetylation and
molecular weight of chitosan prepared were also compared. The chitosan
samples prepared from lactic acid treated showed high viscosity than HCI treated
samples. But degree of deacetylation is more in HCI treated samples than lactic
acid treated ones. Characteristics of protein liquor obtained like its biogenic
composition, amino acid composition, total volatile base nitrogen, alpha amino
nitrogen etc. also were studied to find out its suitability as animal feed
supplement.Optimization of fermentation parameters for Lactobacillus brevis
fermentation study was also conducted and parameters were standardized. Then
detailed fermentation study was done in 20%wlv jaggery broth for 17 days. Also
the effect of two different acid treatments (mild HCI and lactic acid) used for initial
pH adjustment on chitin production were also studied. In this study also trend of
changes in pH. changes in sugar concentration ,microbial count changes were
similar to Lactobacillus plantarum studies. At the end of fermentation, residual
protein in the samples were only 32.48% in HCI treated samples and 31.85% in
lactic acid treated samples. The residual ash content was about 33.68% in HCI
treated ones and 32.52% in lactic acid treated ones. The fermented residue was
converted to chitin with good characteristics by treatment with 1.2MNaOH and
1NHCI.Characteristics of chitin samples prepared were studied and extent of Nacetylation
was about 84% in HCI treated chitin and 85%in lactic acid treated
ones assessed from FTIR spectrum. Chitosan was prepared from these samples
by usual chemical method and its extent of solubility, degree of deacetylation,
viscosity and molecular weight etc were studied. The values of viscosity and
molecular weight of the samples prepared were comparatively less than the
chitosan prepared by Lactobacillus plantarum fermentation. Characteristics of protein liquor obtained were analyzed to determine its quality and is suitability as
animal feed supplement.Another strain used for the study was Bacillus subtilis and fermentation
was carried out in 20%w/v jaggery broth for 15 days. It was found that Bacillus
subtilis was more efficient than other Lactobacillus species for deprotenisation
and demineralization. This was mainly due to the difference in the proteolytic
nature of the strains. About 84% of protein and 72% of ash were removed at the
end of fermentation. Considering the statistical significance (P<O.05) in the
extent of demineralization and deproteinisation, we have taken 0.8N HCI for the
demineralization study and 0.6M NaOH for deprotenisation study. Properties of
chitin and chitosan prepared were analyzed and studied. |