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PROFORMA (available in DBT website)
PROFORMA FOR SUBMISSION OF PROJECT PROPOSALS ON RESEARCH
AND
DEVELOPMENT, PROGRAMME SUPPORT
(To be filled by the applicant)
PART I: GENERAL INFORMATION
1. Name of the Institute/University/Organisation
submitting the Project Proposal : SASTRAUNIVERSITY
2. State : Tamilnadu
3. Status of the Institute : Deemed University
(Please see Annexure-I)(Project Performa downloads in DBT site, Sl. No 20)
4. Name and designation of the Executive Authority
of the Institute/University forwarding the application : The Registrar
SASTRA University
5. Project Title : Bioremediation of Shrimp Bio waste
By Using Natural Probiotic for the
production of Chitinase and Astaxanthin
6. Category of the Project (Please tick): R&D
7. Specific Area : Environmental and industrial
8. Duration : 3 years
9. Total Cost (Rs.) : 42,10,600
10. Is the project Single Institutional or
Multiple-Institutional (S/M)? : Single
11. If the project is multi-institutional, please furnish the following : Nil
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12. Scope of application indicating anticipated product and processes
Shrimp processing is one of the major food industries in India. Shrimp processing
waste is the single largest industrial fish waste in the country causing diverse
environmental problems. Only 40% of the shrimp is edible and remaining 60 % account
for the processing discards. It is estimated that organisms annually produce from 100 to
200 billion tons of this “shell material”. This constitutes an enormous quantity of unused
biomass. These discards find very little practical application at present and are
categorized as a major environmental contaminant. Effective utilization of the waste can
resolve many of the environmental concerns facing the shellfish processors. In addition to
the traditional uses, shrimp waste is one of the important natural sources of carotenoid
and chitin.
PHASE-I(Extraction of Astaxanthin)
Astaxanthin (3,30-dihydroxy-b,b-carotene- 4,40-dione) is one of the most
important pigments in aquacultural industry to provide a desirable reddish-orange color .
In addition, antioxidant properties of astaxanthin have been reported to surpass those of β-
carotene or even α-tocopherol.Due to its outstanding antioxidant activity, astaxanthin has
been attributed with extraordinary potential for protecting organisms against a wide range
of ailments such as cardiovascular problems, different types of cancer and some diseases
of the immunological system.Different methods were used to extract astaxanthin such as
fermentation, enzymatic, and organic solvent processes. Organic solvents such as acetone,
methanol, isopropyl alcohl, petroleum ether have been used to extract astaxanthin from
crustacean byproducts.
PHASE-II(Use of Crude Shrimp Shell Powder for Chitinase Production)
Treatment of this biowaste using hazardous chemicals involves alkalis (usually
4% NaOH) for deproteinization and strong acids (4% HCl and acetic acid) for
demineralization, making this process ecologically aggressive and is a source of
pollution.
Chitin is the second most abundant polysaccharide in nature, after cellulose, and it
largely exists in wastes from the processing of marine food products (crab, shrimp and
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krill shells). About 1011 t of chitin is produced annually in the aquatic biosphere alone.
The waste generated from the worldwide production and processing of shellfish is a
serious problem of growing magnitude. This abundant waste may pose environmental
hazard due to the easy deterioration.
Production of microbial chitinase is one of the primary economic in the utilization
of shrimp processing solid waste. In recent years a considerable interest has been made in
using shrimp processing waste for chitinase production by solid state fermentation.
Production of microbial chitinolytic enzymes has received much attention as one step in a
bioconversion process to treat shellfish waste chitin and its application in the production
of Nacetyl- D-glucosamine for use in food and pharmaceutical.
13. Project Summary (Not to exceed one page. Please use separate sheet).
Shrimp waste is one of the important sources of chitin and natural carotenoids.
Shrimp biowaste being alkaline (pH 7.5-8.0) supports the growth of undesirable
putrefying micro flora resulting in spoilage. The waste generated from the world-wide
production and processing of shellfish is a serious problem of growing magnitude. This
abundant waste may pose environmental hazard due to the easy deterioration. Hence this
project focuses on using the dried shrimp shellsin two phases. One is the extraction of
ASTAXANTHINfrom the shells, a red orange pigment and an powerful anti-oxidant
which has vast application in aquaculture, cosmetic, and functional foods. There are also
potential markets for natural carotenoids in soft drinks, ice cream, desserts, candies, meat
products and pet and aquaculture feeds. Other one is microbial production of
CHITINASE enzymes using dried shrimp shell powder as substrate by
fermentation.There is an increasing interest in the use of chitinases for the control of
moulds, insects, nematodes, and production of different chitin oligomers. The ability of
chitinase to hydrolyze chitin makes it very useful for the production of value added
products suchas sweeteners, growth factors, and single cell protein. The exploitation of
chitinase with respect to plant defense can be done a variety of ways. The enzyme can
also be used in free or immobilized form to kill fungi and insects in affected areas.
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PART II: PARTICULARS OF INVESTIGATORS
(One or more co-investigators are preferred in every project. Inclusion of co-
investigator(s) is mandatory for investigators retiring before completion of the project)
Principal Investigator:
14. Name : Dr. N.Mahesh
Date of Birth : 06.07.1979 Sex (M/F) : Male
Designation : Assistant Professor – III
Department : Department of Chemistry and Biosciences
School of Chemical and Biotechnology
Institute/University: SASTRA UNIVERSITY
Address : Srinivasa Ramanujan Centre,
Kumbakonam, Thanjavur (Dt), Tamilnadu
PIN : 612 001
Telephone : 914352426823 Fax : 914352402460
E-mail : [email protected], [email protected]
Number of research projects being handled at present : Nil
Co-Investigator
15. Name : Dr. S. Balakumar
Date of Birth : 09.07.1976 Sex (M/F) : Male
Designation : Assistant Professor – III
Department : Department of Chemistry and Biosciences
School of Chemical and Biotechnology
Institute/University: SASTRA UNIVERSITY
Address : Srinivasa Ramanujan Centre,
Kumbakonam, Thanjavur (Dt), Tamilnadu
PIN : 612 001
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Telephone : 914352426823 Fax : 914352402460
E-mail : [email protected]
Number of Research projects being handled at present : 1
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PART III : TECHNICAL DETAILS OF PROJECT
(Under the following heads on separate sheets)
16. Introduction (not to exceed 2 pages or 1000 words)
Shrimp processing is one of the mostimportant marine industries that
generatesconsiderable quantities of shrimpwaste consisting of head, shell and tail ofthe
shrimp. The body parts processed forhuman consumption comprises approximately70%
of the total shrimp landing,so there is a tremendous tonnage ofshrimp waste produced.
The traditional chemical method creates a disposable problem due to large
amounts of toxic waste without further treatment would pollute the environment. As
regulations have become stricter now, there is a need to treat and utilize the waste in most
efficient manner. Therefore there is a significant interest regarding recycling of shrimp
biowaste. Hence the conversion of shrimp biowaste into ensilage advantageously
upgrades the biowaste with this approach being eco-friendly safe, technologically flexible
and economically viable. An alternative method, using fermentation by microorganisms
has been emerged, which can do considerable extent replacing the expensive and non-
environmental friendly chemical process.
Astaxanthin, the main pigment found in crustacean and salmonids provides the
desirable reddish-orange colour in these organisms. In addition to its pigmentation
function, one of the most important properties of astaxanthin is its antioxidant activity. It
has been reported that the antioxidant activity of astaxanthin is 10 times higher than other
carotenoids such as zeaxanthin, lutein, canthaxantin, and β-carotene.Numerous studies
have identified astaxanthin antioxidant mechanisms that quench active oxygen species
and free radicals in vitro and in vivo. Because of its antioxidant properties, astaxanthin
may have a role in the treatment of chronic diseases such as cardiovascular diseases,
cataract development, macular degeneration, and some types of cancer. Astaxanthin is
widely used in aquaculture, cosmetic, and functional foods.
Chitinases, capable of catalyzing the hydrolysis of chitin to its monomer N-acetyl-
D-glucosaminefrom various sources, such as bacteria, fungi, yeast and plants, have been
characterized. The chitin wastesespecially derived from sea-food-processing units
causeremarkable environmental problems worldwide. Chitinasesplay important roles in
industrial applications in the treatment of chitin. The production of inexpensive
chitinolyticenzymes is an important element to the utilization of shellfishwastes, which
not only solves environmental problems but alsohelps to extend the applications of
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microbial chitinases.Chitinases could also be used extensively in biological researchfor
the generation of fungal protoplasts due to their ability todegrade fungal cell wall. The
hydrolytic property of chitinasesmakes them attractive as environmentally safe
biocontrolagents. Using microbial fermentation techniques, chitinase can be produced
from the dried shrimp shell powder.
16.1 Origin of the proposal
Shellfish take a major share in the seafood processing and earn a significant
amount of foreign exchange through export in India. The industries, however, are facing
with severe problem in disposing of the formidable quantity of shellfish solid wastes.
About 40% of the processed seafood consists of shrimp and hence a major share of
shellfish waste composed ofshell and head of the processed shrimp. It can be estimated
that depending on the species and methods of processing waste fraction constitute more
than 50% of the landed shrimp, which amounts an average of 1.5 lakh MT annually. The
conventional method of shellfish processing includes waste disposal by ocean dumping,
incineration and land filling. However, factors such as cost of transportation and
environmental pollution have prompted the need for alternative disposal methods.
Bioconversion into value added products is probably the most cost-effective and
environment friendly method for waste utilization which have several advantages such as
elimination of environmental pollution, improve the profitability of seafood processing
industry, promote the economic value of marine products. A number of possibilities for
bioconversion of shellfish waste have been reported, but each of these has problems of
practicability or economics. In India not much attention has given for the total economic
utilization these solid wastes.
16.2 (a) Rationale of the study supported by cited literature (b) Hypothesis (c) Key
questions.
a. From the various literatures proved that there is an urgent need to find a remedy
for the shrimp biowaste management that is reuse of the shrimp shells. Chemicals
are hazardous and pollution causing in nature. Hence the bio conventional
methods like fermentation are used.To produce certain enzymes like chitinase,
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dried shrimp powder can be used as a substance which is rich in chitin and other
carotenoids.
b. In this proposed study, large amount shrimp shells can be used for the extraction
of astaxanthin which is powerful anti-oxidant by chemical extraction method, and
the production of chitinase enzyme used this shell powder as a substrate.
c. Though many methods are used for the management shrimp shell wastage, they
are pollution causing in nature. The proposed methods for the reuse of shells are
ecofriendly and low expensive.
16.5 Current status of research and development in the subject (both international
and national status)
National
S.Sindhu (2011)suggested that, extraction of carotenoids from the shell waste of
the Arabian red shrimp Aristeusalcockiwas investigated using different organic solvents,
and vegetable oils, under wet and dry conditions, with and without deproteinization. The
highest carotenoid yield was obtained with non-deproteinized wet waste extracted using
acetone. The carotenoid yield was found to be double that of Pandalus borealis shell
waste, which is currently used as one of the commercial sources of natural astaxanthin.
KandraPrameela et al studied about the production of chitin and carotenoids from
shrimp biowaste by using naturalprobiotic. The fermentation studies were determined by
varying inoculumlevels from 1 to 10%, glucose concentration from 0 to 15% and
incubation time 0 to 72 hours. As the process by using natural probioticwas found to be
efficient, economical, it becomes an alternative method for hazardous chemicalmethod
used in extraction of chitin and carotenoids.
S. Sindhu and P.M. Sherief, Extraction, Characterization, Antioxidant and Anti-
Inflammatory Properties of Carotenoids from the Shell Waste of Arabian Red Shrimp
Aristeusalcocki, Ramadan 1938, The Open Conference Proceedings Journal, 2011, 2, 95-
103 95.
KandraPrameela, Ch. Murali Mohan, P.V. Smitha, K.P.J. Hemalatha, Bioremediation Of
Shrimp Biowaste By Using Natural Probiotic For Chitin And Carotenoid Production An
Alternative Method To Hazardous Chemical Method, International Journal of Applied
Biology and Pharmaceutical Technology, Volume: I: Issue-3: Nov-Dec -2010.
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International
San-Lang wang et al .,worked on the production of chitosanase and a protease
were purified from the culture supernatant of Serratia sp.TKU016 with shrimp shell as the
sole carbon/nitrogen source. The molecular masses of the chitosanase and protease
determined by SDS–PAGE were approximately 65 and 53 kDa, respectively.
SwiontekBrzezinska et al.,examined the level of respiration activity of water and
sediment microorganisms in the presenceof shrimp waste. Results demonstrate that the
number of heterotrophic bacteria and fungi in water and bottom sediments were variable.
The analyzed groups of microorganisms predominated in bottom sediments with the
number of heterotrophic bacteria significantly exceeding that of fungi. The proportion of
microorganisms capable of decomposing chitin was greater among fungi than among
heterotrophic bacteria.
Po-Min Kao et al., concluded that the feasibility of using membrane mode
fermentation operations for the continuous chitinase production by Paenibacillus sp.CHE-
N1 .The bioreactor with a membrane outer recycling loop was used to evaluate the effect
of membrane pore size on cell retention efficiency, permeate flow rate, fouling, and
chitinase recovery in permeate. The results showed that at a transmembrane pressure of
0.9 kg/cm2, M 9 microfiltration column with a nominal pore size of 300 kDa exhibited
the best microfiltration characteristics and was used for themembrane mode operation.
San-Lang Wang , Tao-Jen Chang ,Tzu-Wen Liang,Conversion and degradation of shellfish
wastes by Serratia sp. TKU016 fermentation for the production of enzymes and bioactive
materials, Biodegradation (2010) 21:321–333
M. SwiontekBrzezinska, E. Lalke-Porczyk, W. Donderski,Utilization of Shrimp Waste as a
Respiration Substrate by Planktonic and Benthic Microorganisms, Polish J. of Environ.
Stud. Vol. 17, No. 2 (2008), 273-282
Po-Min Kao, Shu-Chen Huang, Yung-Chi Chang, Yung-Chuan Liu, Development of
continuous chitinase production process in a membrane bioreactor by Paenibacillus sp.
CHE-N1, Process Biochemistry 42 (2007) 606–611.
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16.6 The relevance and expected outcome of the proposed study
1. Utilizing large number of shrimp shells for the production of the enzyme and
extraction of bioactive compound.
2. The enzyme, chitinase has more importance in the field of food and pharmaceutical
which is going to be produced.
3. Astaxanthin, a powerful anti-oxidant and reddish orange pigment which has widely
used in aquaculture, cosmetic, and functional foods.
4. We aim at achieving an effective, safe and economical solution for the management of
shrimp shell waste.
16.7 Preliminary work done so far
The P.I has collected sufficient Literature on the above research area and has
worked in the production of different enzymes using different substrates. Moreover, four
M.Phil students were worked and six PG Students are working under the P.I focusing on
the bioactive compound extraction and fermentation techniques.
17. Specific objectives (should be written in bulleted form, a short paragraph
indicating the methods to be followed for achieving the objective and verifiable
indicators of progress should follow each specific objective)
To collect Shrimp waste from shrimp processing landing centers at Poombukar
and Pondicherry, South East costal region of Tamilnadu.
PHASE-I
To extract Astaxanthin by chemical methods.
Isolation of Lactobacillus plantarum and Lactobacillus acidophilus from shrimp
and vegetable waste.
To develop the fermentation process to extract Astaxanthin esters and other
bioactive compounds.
To optimize the inoculum size, substrate level and physiological condition.
To analyze the statistical the response variables.
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To study the qualitative and quantitative estimation of compounds.
PHASE-II
a) To partially characterize crude chitinase produced by Bacilluslicheniformisin batch
culture.
b) To study optimum condition for enzymatic degradation of pretreated shrimp waste
using Central Composite Design.
c) To optimize factors those include shrimp waste, initial pH and temperature.
18. Work Plan: should not exceed 3-4 pages (the section can be divided according to
the specific aims and under each specific aim; the following should be stated clearly
as sub headings)
18.1 Work plan (methodology/experimental design to accomplish the stated aim)
PHASE-I Extraction and production of Astaxanthine
Microorganism and Culture Media
Lactobacillus plantarum and Lactobacillus acidophilus will be isolated from the
Shrimp waste region and food contaminants.
Preparation of Shrimp Waste
Shrimp waste comprising of head and carapace will be collected from a shrimp
processing landing centers at Poombukar and Pondicherry, South East costal region of
Tamilnadu and will be transferred to laboratory with desired condition. The dried shell
will be weighed and homogenized in a laboratory mixer.
Extraction of Astaxanthin
From the Shrimp waste powder homogenate is extracted with hexane and glass
bead and vortex for a minute and placed in water bath for 10 min. Aqueous layer will be
separated by 3000 rpm for 5 min until the hexane is colorless the step will be repeated.
Finally DMSO will be added and vortex vigorously and place in water bath to separate
carotenoid.
Microbial extraction of Astaxanthin in fermentation medium
In the MRS broth the Lactobacillus cultures will be inoculated with shrimp waste
homogenate as substrate and incubation will be for 3 days at 30°C, in the presence of 5%
CO2. The fermentative culture medium will be extracted with respective solvents and
Astaxanthin will be extracted.
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Optimization of conditions for carotenoid extraction
The condition will be optimized with the effect of combination process variables
such as lactose sugar, yeast extract and other physical modification on the medium and
condition and their interactions on the response variables will be measured.
Purification of Astaxanthin
The concentrated extract from the methods will be subjected to
Spectrophotometer, TLC, HPLC, IR and NMR to quantify the amount of Astaxanthin.
Production of bioactive compounds
For the production of bioactive compounds the first step is to isolate and screen
the cultures shows any bioactive molecules.
Optimizing Fermentation condition
The condition will be optimized with the effect of combination process variables
such as sugar, yeast extract and other physical modification such as temperature, pH and
inoculum size and incubation time on the medium and condition and their interactions on
the response variables will be measured.
Enzyme and Protein Assay
Using appropriate solvent enzymes and proteins will be extracted and analyzed
with spectrophotometer.
Analysis of compounds
The concentrated extract from the methods will be subjected to TLC, HPLC, IR
and NMR to quantify the amount of compounds.
PHASE-II Production of Chitinase
Microorganism
Bacillus sp is a thermophilic organism with the ability to grow in extreme
temperature around 75°C and lively stable in temperature ranging from 30 to 80°C.
Sakai et al. (1998) described pH 6.5 and pH 6.0-9.0 is identified to be theiroptimum and
stable pH for the production of chitinase. Bacillus licheniformis strain will be isolated
from shrimp and vegetable waste,used in the productionof crude chitinase.
Preparation of Inoculum
Preparation of inoculum medium will be performed using Hungate technique. It
will be carried out by transferring pure bacterial colonies from solid to liquid RGM using
sterile loop under aseptic condition. The medium is sparged with a mixture of N2:CO2
(80:20) gases to eliminate present oxygen gas. The medium is prepared in serum bottle
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with crimped metal seal in order to ensure an anaerobic condition for the bacterial
growth. Following inoculating, head-space of the serum bottle is sparged with oxygen
free gas in sterile condition to remove residual oxygen. The inoculum was then incubated
at 50°C until OD550nm reached 0.6-0.8 before transferring 10% (v/v) of inoculum into
Schott bottle containing fermentation medium for batch process.
Microbial production of chitinase in fermentation medium
In the RGM broth the Bacillus cultures will be inoculated with shrimp waste
homogenate as substrate and incubation will be for 3 days at 30°C, in the presence of 5%
CO2.
Identification of pH Optimum and Stability
The optimum pH will be determined by measuring chitinase assay (DNS method)
with various pH values ranging from 3 to 9. The study for pH stability will be determined
by prolonging incubation period from one to two hours at various pH values before
performing chitinase assay.
Identification of Temperature Optimum and Stability
The optimum temperature will be determined by measuring chitinase assay (DNS
method) in 0.2 M citrate-phosphate buffer pH 6.0 for 1 hour at temperature ranging from
30°C-70°C. The stability of chitinase towards temperature will be measured by
prolonging incubation period for 1 to 2 hours in 0.2 M citrate-phosphate buffer pH 6.0 at
temperatures ranging from at 30°C to 70°C before performing chitinase assay.
Molecular Weight Determination
The purpose of SDS-PAGE is to separate proteins according to their
electrophoretic mobility which is a function of length of polypeptide chain or molecular
weight as well as higher order protein folding, post-translational modifications and other
factors. This method will be used to compare and characterize the different separated
proteins.
Analytical Method
During fermentation process, 10 mL of batch culture sample will be taken atdaily
intervals for every 2 hours for 24 hours for protein assay. Sample taken will be
centrifuged at 4000 rpm for 15 minutes at 4°C. The supernatant was used todetermine
chitinase activity and protein concentration whilst the pellet is resuspended using distilled
water and used in determining the cell biomass.
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Purification of chitinase
Purification of chitinases will be done by DEAE-Sepharose CL-6B
chromatography.
18.2 Connectivity of the participating institutions and investigators (in case of multi-
institutional projects only)
18.3 Alternate strategies (if the proposed experimental design or method does not
work what is the alternate strategy)
The present work aims to find an alternative to utilize the shrimp shells for the
production of chitinase enzyme and the extraction of astaxanthin. Unfortunately, if it fails
other bioactive compounds like caroteinoids will be produced which has vast application
in the food and pharmaceutical field.
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19. Timelines: (Please provide quantifiable outputs)
Work Elements
3 Years Duration
I Year II Year III Year
A B C D A B C D A B C D
Literature review &
Bibliography documentation
Systematization of project studies
Purchase of Equipments
Collection & isolation of
microorganism from Shrimp waste
Screening of microorganisms
Phase – I and II
Chemical extraction of astaxanthine
Isolation & Identification of
Microorganism
Shake flask fermentation studies for
substrate level and environmental
conditions
Studies using fermenter
1.Media Optimization
2.Fermentation & Recovery process
Identification & Characterization of
compounds
Compiling of data & preparation of
final report
Patent filing if new bioactive
compound
Fig.1 Panoramic presentation of various time schedules related to the project
Note : Each column marked as A, B, C and D indicates a period of three months
(we regret the unequal size)
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20.Name and address of 5 experts in the field
Sr.No. Name Designation Address
1. Prof. M. Lakshmanan
Former Vice –
Chancellor,
MaduraiKamarajUniversity,
Madurai, Tamilnadu, India
2. Dr. T.Thirunalasundari Professor Dept. of Biotechnology,
BharathidasanUniversity,
Thiruchirapalli, tamilnadu
3.
Dr.A.p. Liton Principal Scientist
and Incharge
CMRI, Marine biotechnology
lab, Vizhinajm research centre
695 521, Kerala
4. Dr. A. Palavesam Head Centre forMarine Science &
Technology
Rajakkamangalam
Kanyakumari District
Tamil Nadu.
629 502
5. Dr.K.Singaravadivel Professor & HOD Indian Institute of Crop
Processing Technology
Ministry of Food Processing
Industries, Government of India,
Pudukkottai Road,
Thanjavur - 613 005
Tamil Nadu. India
Email ID : [email protected]
Mobile No. : +91-9750968405
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PART IV: BUDGET PARTICULARS
Budget (In Rupees)
A. Non-Recurring (e.g. equipments, accessories, etc.)
S. No. Item Year 1 Year 2 Year 3 Total
1.
2.
3.
Fermenter with Accessories
Environmental rotatory shaker
Rotary vaccum Evoporator
12,50,000
3,00,000
2,00,000
12,50,000
3,00000
2,00,000
Sub-Total (A) =17,50,000
B. Recurring
B.1 Manpower (See guidelines at Annexure-III)
S. No. Position
No.
Consolidated
Emolument
Year 1 Year 2 Year 3 Total
1.
Junior Research
Fellow (1 No.)
(M.Sc) @ 1
15600 @ 24
month and
18200@12
month
inclusive of
30% HRA
374400 374400 436800
11,85,600
Sub-Total (B.1) = 11,85,600
B.2 Consumables
S. No. Item
Quantity Year 1 Year 2 Year 3 Total
1. Consumables/
Chemicals
As Required 3,50,000 3,50,000 3,50,000 10,50,000
Sub-Total (B.2) = 10,50,000
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Other items Consolidated
Emolument
Year 1 Year 2 Year 3 Total
B.3 Travel
25000 25000 15000 65000
B.4 Contingency
50000 50000 60000 160000
Sub-total of B
(B.1+B.2+B.3+B.4)
799400 799400
861800 2460600
Grand Total (A + B) 17,50,000 24,60,600 42,10,600
Note : Please give justification for each head and sub-head separately
mentioned in the above table.
JUSTIFICATION FOR EQUIPMENT
Fermenter with Accessories
This instrument will be useful for the production of antibiotics. Fermenter have
sophisticated facilities and optimized condition support to enhance the production.
Moreover this study confines with growth kinetics of the organism and their metabolic
products, it is must for the conditions very much warranted for the successful completion
of the project. Naturally biomolecules will produce only limited amount by the microbial
cells so it requires bulk medium for the large scale production. We have fermenter for the
regular usage of B.Tech and M.Tech biotechnology students, hence it will make difficult
to complete our research. Moreover this instrument is available in thanjavur campus, but
our proposed project to be held at kumbakonam it takes one hour travel to do our work.
Our proposed work requires continue usage of this instrument, to mitigate this we require
additionally one more to cope up our research successfully.
Environmental rotatory shaker
This instrument is essential to study the shake flask level fermentation in a
controlled environmental condition. So, that the enzyme yield and screening of
optimization can be performed. Before going to pilot study using fermenter the
parameters can be optimized by shake flask level minimize the substrate level and time is
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concern. Moreover, it contains the ambient temperature controller that could monitor the
environmental temperature.
Rotary Vaccum Evaporator
As the proposed work requires chemical extraction for Astaxanthin, we require
large amount of solvents for the extraction process. To reduce the cost of the solvent and
usage we require rotary evaporator for materialize the minimum amount of chemicals.
Manpower
Junior Research Fellow (M.Sc)
As we are going to carry out the Isolation of Microorganism and their biochemical
& molecular characterization, Fermentation process and characterization of biomolecules
it leads to a large quantum of experimental requirements in the field of Microbiology and
bio-chemical studies. In view of that, supportive analytical persons in the laboratory are
necessary. The laboratory backup on experimentation will be supported by one J.R.F
having knowledge in the fields of Microbiology /Bio-Chemistry/ Biotechnology.
Consumables /Chemicals
Consumables / Chemicals cost includes particularly chemicals, culture media,
accessories and glasswares. Taking into consideration the cost of Biochemical
compounds and the various tests used for the different experiments to be carried out using
Borosilglasswares the requirement of Rs.3, 50,000 per year is reasonable. Over and
above, microorganisms have to be procured, isolated and to be maintained. Also Standard
antibiotics have to be procured, maintained, and their effectiveness to be tested. The cost
of animals and the maintenance of the same warrant an appreciable sum in this projection.
Contingency (Books/Journals)
In spite of having the basic books in our library, for our ready reference in the
laboratory, we have to keep a set of books / recent publications. Keeping in to account the
high cost of recent books in Microbiology, Biochemistry, Fermentation technology and
downstream processing a reasonable budget is being asked here.
Recurring
The PI, Co-PI and other project staff have to go for seminars and workshops. Such
activities expect a large Chunk of T.A. / D.A. from this allocation. Nevertheless, our
University research guidelines expects the scholars who have registered for Ph.D., must
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publish 2 papers before submission of thesis. So, it warrants project staff to move around
for the participation of seminars, workshops, etc.
Also it is necessary to publish our research findings then and there in journals,
which require postage, stationary, printing, photocopying, etc. Over and above, the
maintenance of the existing instruments, spare parts for the instruments, up-keep of the
laboratory, etc., requires an appreciable quantum of expenditure. Moreover, in the third
year, we need an appreciable sum that includes the cost involved in the final report
preparation. In view of that, Rs.1,60,000/- for 3 year recurring contingency is projected
here.
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PART V : EXISTING FACILITIES
Resources and additional information
1. Laboratory:
Lab Particulars Sq.ft
Central Animal Facility 16598
Sophisticated Central Instrumentation Facility 2103
In-vivo and In-vitro Pharmacology lab 1726
Phytochemistry Lab 1872
Biochemistry Lab 1726
Histopathology lab 400
Pharmacoinformatics Lab 878
Optical Room 405
Microbiology Lab 200
Sedimentology Lab 878
Botany Lab and Library 878
Computer Applications Lab 986
Geochemical Lab 2103
M.Pharmacy class Rooms (878 sq.ft x 4 Nos.) 3512
Administrative Area 878
Pharmaceutics Lab 3816
Chemicals and Glasswares Stock Room 782
Staff Room 2611
NMR Room 495
a. Manpower - 95
b. Equipments
22
State–of –the–art facilities in SASTRAUNIVERSITY
To accomplish the above objectives, the Centre is well equipped with the state-of –the-
art facilities in SASTRA such as:
Name of the Instruments Supplier
NMR (300MHz) Bruker
Field Emission Scanning Electron Microscope with
energy dispersive spectros
Joel
GC-MS with FID & FPD Perkin Elmer
GC with ECD & NPD Perkin Elmer
Liquid Scintillation Beta Counter Perkin Elmer
Preparative HPLC Waters
HPTLC CAMAG
CHN/S /O Perkin Elmer
Gamma Counter Perkin Elmer
Fluorescence Spectrophotometer (F-7000) Hitachi
Fully Automated Chemistry Analyzer Merck
FTIR Perkin Elmer
AAS Perkin Elmer
Graphite Furnace Perkin Elmer
Hematology 5 part Analyzer Trivitron
Particle Size Analyser AccuSizer
Polarimeter Rudolf
APO Zoom Stereomicroscope & Digital camera Leica
Polarized Research Microscope with camera Leica
Trinocular Light Microscope with camera Olympus
PCR Eppendorf
Fermentor New Brunswick Scientific Company
Laminar airflow chamber New Brunswick Scientific Company
Lyophilizer Martin Christ
Multiwave Digestion System Anton Paar
23
Ultra Microtome Leica
Ultra Pure Water Purification System (ELIX-3 &
MILLI @ GRADIENT)
Millipore
UV Spectrophotometer Perkin Elmer
Embedding machine Leica
Tissue stainer Leica
Refrigerated Centrifuge Sigma
Western Blot Unit Biorad
ELISA Reader Qualigens
ELISA Washer Qualigens
Semi –auto analyzer Qualigens
Deep Freezer Cryo Scientific
Analytical Balance Metlar
Flame Photometer (2 Nos.) Systronics
Gama Ray Counter Thermo
Fume cupboard Klean Zone Systems
Electrophoresis unit Biotech
Cooling Centrifuge Remi Motors
Weighing Machines – 5 Nos. Schmadzu.
Eddy’s hot plate INCO
De-Ionizer Ion Exchange Limited
Student Universal Kymograph INCO
Histamine chamber INCO
Dale’s mono bath INCO
Tele-thermometer INCO
Water Kit Analyzer Systronics
Minor equipments like, Muffle Furnace, pH Meter
(digital), Rock grinder with agate pestle and mortar
Aravind Scientific
Rotary Vacuum Evaporator Super fit
Plethysmograph Madras Scientific Co., Trichy
Rota Rod Apparatus INCO
Autoclave (3 Nos) Yorco& Reliance Instruments
24
Hot Air Ovens Semco
Shaker Scigenics Biotech
Digital Colony Counter Deep Vision
Centrifuges Remi
Cyclomixer RemiEquipments Pvt. Ltd.,
Deep Freezer Blue Star
Incubators Semco
UV Spectrophotometer (2 Nos ) Elico
Magnetic Stirrers RAAGAA Industries
Cooling Centrifuge Remi
BOD incubator Aravind Scientific Suppliers
Laminar Airflow Chamber Kleanzone System
Inverted Tissue Culture Microscope Weswox
Stereo Zooming Microscope Weswox
Binocular Microscope Weswox
Student Microscope Ajay Optics &Weswox
Fast Protein Liquid Chromatography Akta Prime
Membrane Filteration Mid Gee
2.Other resources such as clinical material, animal house facility, glass house.
Experimental garden, pilot plant facility etc.
We have sophisticated Air-Conditioned Central Animal Facilityof 16598sq.ft with
CPCSEA clearance.
Moreover, the animal house is supported by a list of equipments as shown below:
1. Digital Plethysmometer (Stoelting, USA)
2. Anesthesia System (Stoelting, USA)
3.Hotplate Analgesia Meter (Columbus, USA)
4.Tail Flick Analgesia Meter (Columbus, USA)
5. Electro convulsive device (UGO BASILE, Italy)
6.16 channel Data Acquisition System with Isolated Organ Bath, Lagendroff&
Non-Invasive Blood Pressure System (Biopac Systems, USA)
25
7. Metabolic Cages for mice, rats, guinea pigs & rabbits (Tecniplast, Italy)
8. Animal Operating Table& Heated Blanket systems (Vet Tech, Solutions, UK)
9. Telemetry Transmitter and Receiver system (Data Sciences International, USA)
10. Type II Long IVC System for Mice (Allentown caging equipment, USA)
11. Bio Containment Unitof 42 cages for Mice (Allentown caging equipment, USA)
12. Safety Cabinet 1016 mm W/Hydraulic Lift & UV light (Allentown caging, USA)
26
PART VI: DECLARATION/CERTIFICATION
It is certified that
a) the research work proposed in the scheme/project does not in any way duplicate the
work already done or being carried out elsewhere on the subject.
b) the same project proposal has not been submitted to any other agency for financial
support.
c) the emoluments for the manpower proposed are those admissible to persons of
corresponding status employed in the institute/university or as per the Ministry of
Science & Technology guidelines (Annexure-III)
d) necessary provision for the scheme/project will be made in the
Institute/University/State budget in anticipation of the sanction of the scheme/project.
e) if the project involves the utilisation of genetically engineered organisms, we agree to
submit an application through our Institutional Biosafety Committee. We also declare
that while conducting experiments, the Biosafety Guidelines of the Department of
Biotechnology would be followed in toto.
f) if the project involves field trials/experiments/exchange of specimens, etc. we will
ensure that ethical clearances would be taken from concerned ethical
Committees/Competent authorities and the same would be conveyed to the
Department of Biotechnology before implementing the project.
g) it is agreed that any research outcome or intellectual property right(s) on the
invention(s) arising out of the project shall be taken in accordance with the
instructions issued with the approval of the Ministry of Finance, Department of
Expenditure, as contained in Annexure-V.
h) we agree to accept the terms and conditions as enclosed in Annexure-IV. The same is
signed and enclosed.
27
i) the institute/university agrees that the equipment, other basic facilities and such other
administrative facilities as per terms and conditions of the grant will be extended to
investigator(s) throughout the duration of the project.
j) the Institute assumes to undertake the financial and other management responsibilities
of the project.
Signature of Executive
of
Institute/University with seal
Date :
Signature of Principal Investigator :
Date :
Signature of Co-Investigator
Date :
28
PART VII: PROFORMA FOR BIOGRAPHICAL SKETCH OF
INVESTIGATORS
Provide the following information for the key personnel in the order listed on PART II.
Follow this format for each person. DO NOT EXCEED THREE PAGES
Name : Dr.N.Mahesh
Designation : Assistant Professor - III :
Department/Institute/University : Department of Chemistry and Biosciences
School of Chemical and Biotechnology
Srinivasa Ramanujan Centre
SASTRA UNIVERSITY
Kumbakonam, Thanjavur (Dt), Tamilnadu
Date of Birth : 06.07.1979 . Sex (M/F) : Male SC/ST : -
Education (Post-Graduation onwards & Professional Career)
Sl No. Institution
Place
Degree
Awarded
Year Field of Study
1. V. H.N.S.N.College M.Sc 2001 Microbiology
2. SASTRA
UNIVERSITY
Ph.D 2008 Microbiology
A. Position and Honors
Position and Employment (Starting with the most recent employment)
Sl No. Institution
Place
Position From (Date) To (date)
1. SASTRA UNIVERSITY Assistant Professor – III /
Scientific Officer
03.05.2006 Till date
2. SASTRA UNIVERSITY Lecturer 15.12. 2004 02.05.2006
3. KISSCO Foods Pvt., Ltd., Microbiologist April 01 April 02
29
Honors/Awards : Nil
Professional Experience and Training relevant to the Project
Two years
B. Publications (Numbers only) .... 9
Books : .................... Research Papers, Reports : ................General articles
:...........................
Patents : .........................Others (Please specify) :..............................................................
Selected peer-reviewed publications (Ten best publications in chronological order)
Jayapradha Ramakrishnan, Murugesh Shunmugasundaram, Mahesh
Narayanan. Streptomyces sp. SCBT isolated from rhizosphere soil of
medicinal plants is antagonistic to pathogenic bacteria. Iranian journal of
biotechnology. 7 (2), 2009
R. Jayapradha , S. Murugesh , N. Mahesh and D. Brahatheeswaran.
Prevalence of ESBL producing strains in Tuberculosis patients. Research
Journal of Microbiology 2007, 2 (5), pages 491 – 495.
Murugesh S, Manivannan S, Mahesh N, Mohanasrinivasan V, Subathara
devi. Bio-solubilization of lignite by marine soil microbes. Indian Journal of
Environmental Protection Volume 23, Issue 10, October 2003, Pages 1081-1092
Mahesh N and Brahateewaran. Anti-hyperglycemic and Anti-hyperlipidemic
activities of aqueous and ehtanolic extract of Cynodon dactylon (Linn) in
streptozotocin induced diabetic rats. Asian Journal of Biochemistry. 2006, 2
(1), pages 66-72
N. Mahesh , S. Murugesh and V. Mohana Srinivasan. Determination of the
presence of Biosurfactant produced by the Bacteria present in the soil
samples. Research Journal of Microbiology, 2006. 1 (4) : 2006 pages: 339-345.
30
C. David Raj , M. Mohamed Shabi , D. Brahatheeswaran and N. Mahesh.
Anti-inflammatory activity of Tylophora indica in albino rats. Journal of
Pharmacology and Toxicology, 2006, 1 (5), pages 490 -492.
Murugesh S, Manivannan S, Mahesh N, Mohanasrinivasan V, Subathara
devi. Production and partial characterization of alkaline protease from
Aspergillus sp. Indian .J. Appl. Micribiol.2003.3 (1) pp:33-35.
Murugesh S, Mohanasrinivasan V, Subathara devi S, Mahesh N,
Manivannan S. Bio-preservation using bacteriocin Nisin. Indian. J. Appl.
Micribiol.2003. 3(1) 23-26.
Communicated and Accepted
Vivek Rangarajan, Mahesh Narayanan, Preamnath Shanmugam. Optmization of
xanthan production by Xanthomonas campestris -NCIM 2961 using Response
Surface methodology, International Journal of Applied Biotechnology and
Biochemistry. 2011 (Accepted)
Narayanan Mahesh, Srinivasan Balakumar, J. Thangam, Arunadevi Ayyadurai,
Rangarajan Vivek. Optimization and enhanced production of cephalosporin c
under solid state fermentation and submerged fermentation from Acremonium
chrysogenum MTCC 431, Journal of Biotechnology. 2011 (Communicated)
Narayanan Mahesh, Srinivasan Balakumar, R. Parkavi, Arunadevi Ayyadurai,
Rangarajan Vivek. Optimization and production of Hyaluranidase by
Streptococcus mitis MTCC 2695., Journal of Biotechnology.2011 Biomolecules
(Communicated)
Narayanan Mahesh, Rangarajan Vivek, Srinivasan Balakumar, Arunadevi Ayyadurai
& Ravichandran. Statistical designing of Enriched pectin extract medium for the
enhanced production of pectinase by Aspergillus niger. Brazilian Journal of
Biotechnology. 2011 (Communicated)
Narayanan Mahesh, Srinivasan Balakumar, P.Indumathi, Arunadevi Ayyadurai,
31
Rangarajan Vivek. Production and optimization of Mevastatin using Penicillium
citrinum NCIM 768, Journal of Microbial and Biochemical technology. 2011
(Communicated)
List maximum of five recent publications relevant to the proposed area of
work.
Vivek Rangarajan, Mahesh Narayanan, Preamnath Shanmugam.
Optmization of xanthan production by Xanthomonas campestris -NCIM
2961 using Response Surface methodology, International Journal of Applied
Biotechnology and Biochemistry 2011 (Accepted)
C. Research Support
Ongoing Research Projects : Nil
Completed Research Projects : Nil
(State only major projects of last 3 years)
Place : Signature of Investigator
Date :
32
PART VII: PROFORMA FOR BIOGRAPHICAL SKETCH OF
CO-INVESTIGATORS
Provide the following information for the key personnel in the order listed on PART II.
Follow this format for each person. DO NOT EXCEED THREE PAGES
Name : Dr.S.BALAKUMAR
Designation : Assistant Professor - III
Department/Institute/University : Department of Chemistry and Biosciences
School of Chemical and Biotechnology
Srinivasa Ramanujan Centre
SASTRAUNIVERSITY
Kumbakonam, Thanjavur (Dt), Tamilnadu
Date of Birth :09.07.1976 . Sex (M/F) : Male SC/ST: ----------
Education (Post-Graduation onwards & Professional Career)
Sl
No.
Institution
Place
Degree
Awarded
Year Field of Study
1. Sivanthi Aditanar College,
Pillayarpuram, Kanyakumari
District
M.Sc 1998 Microbiology
2. BharathidasanUNIVERSITY,
Thiruchirappalli
Ph.D 2008 Microbiology
33
A. Position and Honors
Position and Employment(Starting with the most recent employment)
Sl No. Institution
Place
Position From (Date) To (date)
1. SASTRAUNIVERSITY Assistant Professor – III 14.07.2006 Till date
2. SrimadAndavan college,
Thiruchirappalli
Lecturer 25.06. 1999 13.07.2006
3. Kanchi Sri Krishna
college, Kanchipuram
Microbiologist August, 1998 24.06.1999
Honors/Awards : Received Membership title award from IABMS
Professional Experience and Training relevant to the Project
Two years
B. Publications (Numbers only) ....12
Books : .................... Research Papers, Reports : ................General articles
:...........................
Patents : .........................Others (Please specify) :..............................................................
Selected peer-reviewed publications (Ten best publications in chronological order)
Sivkumar.N, Saravanan.V and Balakumar.S. Bacterial decolourization and
bioremediation of an anaerobically digested molasses spent wash. Asian Journal
of Microbial Biotechnology and Environmental Science, 2006,8(2), 291 – 295.
Sivkumar.N, P. Beulah and Balakumar.S. Ethanol production from Sucrose
medium by an UV mutant Saccharomyces cerevisiae (TBY) Journal of Eco
toxicology and Environmental Monitoring. 2006,16(6), 543 – 547.
34
S. Rajan, S. Balakumar and T. Thirunalasundari. "Antibacerial activity of
Magniferaindica seed kernel on enteropathogenicE.coli" Biomedicine 26: 25-28,
2006.
S. Rajan, S. BalakumarandT. Thirunalasundari. "Antibiotic resistance pattern of
Diarrhoegenic Escherichia coli" Biotechnology: an Indian journal vol.1 (1), 2007.
S. Rajan, S. Balakumarand T. Thirunalasundari. "Antibacterial activity of
Punicagranatum fruit rind of Escherichia coli" Asian Journal of Microbiology,
Biotechnology and Environmental Sciences. Issue 1, 2008.
P.Sampathkumar, B.Dheeba, R.Vinothkannan, S.Balakumar, V.Veena and
A.Shanmughasundaram,. Evaluation of hepatoprotective efficacy of Lippia
nodiflora L. against carbon tetrachloride induced hepatic damage in rats. Journal
of Cell and Tissue research, 8(3): 1595-1598, 2008
S. Sheik Abdullah, M.Arunkumar, S. Balakumar, A. Vijay Anand and T.
Krishnamoorthy. Assessment of phytoconstituents, nutrients and antibacterial
activity of Cardiospermumhalicacabum Linn. Biosciences, Biotechnology
Research Asia. Vol.5(2), 811-814, 2008.
K. Saraswathi and S. Balakumar.Biodecolourization of azodye (pigmented red
208) using bacillus firmus and bacillus laterosporus. Journal of bioscience and
technology. 1(1), 1-7, 2010
Jeyanthi. A., Balakumar.S and Mahalakshmi.T.Incresing bioavailability of
nutrients In fly ash through vermicomposting. J Biosci Tech, Vol 1 (2),2010, 100-
113
Balakumar.S, Mahalakshmi.T Jeyanthi.A. Inhibition of N-Acyl Homoserine
Lactone Mediated Quorum Sensing in Pseudomonas aeruginosa by Phyllanthus
emblica and Quercus infectoria. Journal of Pharmaceutical Sciences and
Research, 2(8), 2010, 521 -526.
Balakumar S, Rajan S , Thirunalasundari T, Jeeva S. Antifungal activity of Aegle
marmelos (L.) Correa (Rutaceae) leaf extract on dermatophytes. Asian Pacific
Journal of Tropical Medicine, 1(4), 2011, 309 – 312.
Balakumar S, Rajan S , Thirunalasundari T, Jeeva S. Antifungal activity of
Ocimum sanctum Linn. (Lamiaceae) on clinically isolated dermatophytic fungi.
Asian Pacific Journal of Tropical Medicine, 4(8), 2011, 654-657.
35
Communicated
Narayanan Mahesh, Srinivasan Balakumar, J. Thangam, Arunadevi Ayyadurai,
Rangarajan Vivek. Optimization and enhanced production of cephalosporin c
under solid state fermentation and submerged fermentation from Acremonium
chrysogenum MTCC 431, Journal of Biotechnology. 2011 (Communicated)
Narayanan Mahesh, Srinivasan Balakumar, R. Parkavi, Arunadevi Ayyadurai,
Rangarajan Vivek. Optimization and production of Hyaluranidase by
Streptococcus mitis MTCC 2695., Journal of Biotechnology.2011 Biomolecules
(Communicated)
Narayanan Mahesh, Rangarajan Vivek, Srinivasan Balakumar, Arunadevi Ayyadurai
& Ravichandran. Statistical designing of Enriched pectin extract medium for the
enhanced production of pectinase by Aspergillus niger. Brazilian Journal of
Biotechnology. 2011 (Communicated)
Narayanan Mahesh, Srinivasan Balakumar, P.Indumathi, Arunadevi Ayyadurai,
Rangarajan Vivek. Production and optimization of Mevastatin using Penicillium
citrinum NCIM 768, Journal of Microbial and Biochemical technology. 2011
(Communicated)
B. Research Support
Ongoing Research Projects : 1
Completed Research Projects : Nil
(State only major projects of last 3 years)
Place : Signature of Co- Investigator
Date :