Open Access Repository of Indian Theses

Ojha, Maheswar

Ribosome Inactivating Proteins (RIPs) are protein or glycoprotein toxins that bring about the arrest of protein synthesis by directly interacting with and inactivating the ribosomes. Such toxins are in general, of plant origin and differ from bacterial toxins that inhibit protein synthesis by mechanisms other than ribosome inactivation. After the toxins had been in the centre of interest in biomedical research for a couple of decades in the end of 19th century, the scientific community largely lost interest in the plant toxins. Interest in these toxins was revived when it was found that they are more toxic to tumor cells when compared to normal cells. Based on their structure RIPs can be classified into three types: Type I RIPs – They consist exclusively of a single RNA-N-glycosidase chain of ~30kDa. Type II RIPs – They consists of chain-A comparable to type I RIPs linked by a disulfide bridge to an unrelated chain-B, which has carbohydrate binding activity. The molecular weight of the type II RIPs is ~60kDa. Type III RIPs – Besides the classical type II RIPs a 60kDa RIP (called JIP60) has been identified in barley (Hordeum vulgare) that consists of chain-A resembling type I RIPs linked to an unrelated chain-B with unknown function.

 In addition to these classes of RIPs there is another group of toxins called four subunit toxins, whose structure is almost similar to type II RIPs, but are made up of two such subunits linked by non-covalent interactions forming tetramers having two A- and two B-chains. The definition and classification of these toxins is not so clear as they are frequently referred to as agglutinins or lectins (e.g Abrus precatorius agglutinins I and II, Ricinus communis agglutinin etc.), having red blood cell (RBC) agglutinating activity. However they have been found to be less toxic and better agglutinins when compared with type II RIPs. 

The present thesis reports the crystal structure of a type II RIP, Abrus precatorius agglutinin-I (APA-I) from the seeds of Abrus precatorius plant. The protein was purified from the plant seed and crystallized. The crystal structure was solved by molecular replacement method. Preliminary crystals of abrus agglutinin were obtained almost thirty years ago and unsuccessful attempts to solve the crystal structure of APA-I were made almost five years ago by other groups. The structure solution of APA-I was obtained at 3.5 Å using synchrotron data set collected at room temperature from a single crystal. Crystal structure is already known for Abrin, another type II RIP isolated from the same seeds. Abrin and APA-I have similar therapeutic indices for the treatment of experimental mice with tumors, but APA-I has much lower toxicity, with lethal dose (LD50) being 5 mg/kg of body weight when compared with Abrin-a (LD50 = 20 g/kg of body weight). The striking difference in the toxicity shown by Abrin and its agglutinin (APA-I) encouraged us to look at the structure function relationship of these proteins, which might prove to be useful in the design and construction of immunotoxins. As apparent from the comparative study, the reduced toxicity of APA-I can be attributed to fewer interactions it can possibly have with the substrate due to the presence of Pro199 at the binding site and not due to any kink formed in the helix due to the presence of proline as reported by the other groups.

In recent years, these plant RIPs which inhibit protein synthesis have become a subject of intense investigation not only because of the possible role played by them in synthesizing immunotoxins that are used in cancer therapy but also because they serve as model system for studying the molecular mechanism of transmembrane translocation of proteins. In silico docking studies were carried out by the author of the thesis in search of inhibitors that could modulate the toxicity of RIPs. Many adenine like ringed compounds were studied in order to identify them as novel inhibitors for Abrin-a molecule and facilitate detailed analysis of protein ligand complex in various ways to ascertain their potential as ligands.

In addition, the candidate has also carried out crystal structural analysis of conformationally constrained, α, β-dehydrophenylalanine containing dipeptides. While there are several studies of molecular self assembly of peptides containing coded amino acids, not much work has been done on molecular self assembly formation utilizing non-coded amino acids. The non-coded amino acid used in our analysis is a member of α, β-dehydroamino acids. These are the derivatives of protein amino acids with a double bond between Cα and Cβ atoms and are represented by a prefix symbol ‘’. They are frequently found in natural peptides of microbial and fungal origins. The presence of α, β-dehydroamino acid residues in bioactive peptides confers altered bioactivity as well as an increased resistance to enzymatic degradation.  Thus, α, β-dehydroamino acid residues, in particular α, β-dehydrophenylalanine (Phe) has become one of the most promising residues in the study of structure-activity relationships of biologically important peptides. The utilization of Phe in the molecular self assembly offers an added benefit in terms of variety and stability. Taking advantage of the conformation constraining property of Phe residue, its incorporation in three dipeptide molecules has been probed. 

In this thesis the crystal structures of the following designed dipeptide are reported. (I). +H3N-Phe-Phe-COO- (FF); (II). +H3N-Val-Phe-COO- (VF); (III). +H3N-Ala-Phe-COO- (AF). The peptides were found to be in the zwitterionic conformation and two (I, II) of the three dipeptides have resulted in tubular structures of dimensions in the nanoscale range.

We have investigated the self-assembly process in a dipeptide incorporating a noncoded, achiral, α ,β-dehydrophenylalanine residue (ΔPhe), an analog of the naturally occurring phenylalanine amino acid with a double bond between Cα and Cβ atoms. Introduction of ΔPhe in peptide sequences is known to induce conformational constraint both in the peptide backbone as well as in the side chain, and to provide the peptide with increased resistance to enzymatic degradation. Here we report the synthesis, characterization, and self-assembly of the dipeptide, Phe–ΔPhe, into distinct tubular structures, which are stable at a broad range of pH conditions and under treatment with proteases.

Abrin and agglutinin-I from the seeds of Abrus precatorius are type II ribosome-inactivating proteins that inhibit protein synthesis in eukaryotic cells. The two toxins share a high degree of sequence similarity; however, agglutinin-I is weaker in its activity. We compared the kinetics of protein synthesis inhibition by abrin and agglutinin-I in two different cell lines and found that ~200–2000-fold higher concentration of agglutinin-I is needed for the same degree of inhibition. Like abrin, agglutinin-I also induced apoptosis in the cells by triggering the intrinsic mitochondrial pathway, although at higher concentrations as compared with abrin. The reason for the decreased toxicity of agglutinin-I became apparent on the analysis of the crystal structure of agglutinin-I obtained by us in comparison with that of the reported structure of abrin. The overall protein folding of agglutinin-I is similar to that of abrin-a with a single disulfide bond holding the toxic A subunit and the lectin-like B-subunit together, constituting a heterodimer. However, there are significant differences in the secondary structural elements, mostly in the A chain. The substitution of Asn-200 in abrin-a with Pro-199 in agglutinin-I seems to be a major cause for the decreased toxicity of agglutinin-I. This perhaps is not a consequence of any kink formation by a proline residue in the helical segment, as reported by others earlier, but due to fewer interactions that proline can possibly have with the bound substrate.

The title compound (systematic name: 3-benzylidene-6-isobutylpiperazine-2,5-dione), C15H18N2O2, an α,β-dehydrophenylalanine containing diketopiperazine, crystallizes in the space group P1 with two molecules in the asymmetric unit arranged antiparallel to one another. The α,β -dehydrophenylalanine (ΔPhe) residue in this cyclic peptide retains its planarity but deviates from the standard conformations observed in its linear analogues. Each type of molecule forms a linear chain with molecules of the same type via pairwise N-HO hydrogen bonds, while weaker C-HO interactions link the chains together to form a three-dimensional network.

A putative homeodomain has been identified in eubacterial genomes, which include several pathogens. The domain is related in sequence to homeodomain, a component specific to transcription factors and playing a very important role in eukaryotes such as controlling the developmental processes of the organism. The putative homeodomain has been characterized utilizing the eukaryotic homeodomain protein sequence signature present in PROSITE as well as the sequence similarity search using BLAST suite for different eubacterial genomes. These findings provide evidence for the occurrence of DNA-binding motif in prokarya similar to that in eukarya.




English

Quantitative Assessment of Gas-Hydrates from Seismic Data

N.T., Manoj

This thesis explores the reactions that lead to polyester formation either by polycondensation, ring opening polymerization or recycling using transesterification reactions, with emphasis on potentials for environmental degradability or reuse.
A method of recycling post-consumer poly(ethylene terephthalate (PET) using transesterification was studied. Shredded flakes of post-consumer PET waste were transesterified with higher diols, such as, 1,4-butanediol, 1,4-cyclohexane dimethanol, and 1,6-hexanediol to yield copolyesters in presence of Ti(iPrO)4 and Sb2O3 as catalysts. The extent of formation of undesirable THF side product was dependent o the molar ratio of PET to 1,4-butanediol and the time of reflux during transesterification. Quantitative insertion of butylene moiety in PET could be achieved under appropriate reaction conditions. Mechanical properties of PBT obtained by transesterification reaction of PET with 1,4-butanediol were comparable to virgin PBT (obtained by direct reaction of dimethylterephathalate with 1,4-butanediol).



Numerical modelling of tidal circulation and studies on salinity distribution in Mandovi and Zuari estuaries

Narvekar, Jayu

Abstract

Seasonal variability of the upper ocean driven by the atmospheric forcing and its regulation of nutrients and chlorophyll in the Bay of Bengal

The upper ocean is the most variable and dynamically active part of the marine
environment that couples the underlying ocean to the atmosphere above through the
transfer of mass, momentum and energy. The thickness of mixed layer is an important
parameter in determining the quantity of heat that is available for exchange with
atmosphere which is capable of triggering several ocean-atmosphere coupled processes.
Mixed layer also plays an important role in determining the chlorophyll biomass and
biological productivity of the upper ocean. The present thesis is an attempt to decipher
the basin-scale variability of the mixed layer in the Bay of Bengal on a seasonal scale and
the factors responsible for it. The aim is also to understand how the changes in water
column nitrate and chlorophyll are linked to the upper ocean variability.

A suite of in situ as well as remote sensing data was used. The in situ data consisted of
temperature, salinity, nitrate and chlorophyll a profiles derived from World Ocean Data
Base and Responsible National Oceanographic Data Centre (RNODC) at National
Institute of Oceanography, which archives the data collected by Indian research ships. In
addition temperature and salinity profiles from Argo were also used. These data were
used to prepare the monthly mean temperature and salinity climatology, while seasonal
climatology was prepared for nitrate and chlorophyll a on a 1ox1o spatial grid. In addition
to this, the high resolution in situ data collected during Bay of Bengal Process Studies
(BOBPS) during summer (6 July to 2 August, 2001), fall intermonsoon (14 September to
12 October, 2002), spring intermonsoon (12 April to 7 May, 2003), and winter (25
November 2005 to 4 January 2006) were also utilized to delineate the effect of mesoscale
variability on mixed layer depth.

As the salinity changed rapidly in the upper layers in the Bay of Bengal due to freshening
by river runoff as well as precipitation, density criteria was used to define mixed layer for
the present study. Mixed layer was defined as the depth at which the density (sigma-t)
exceeds 0.2 kg m-3 from its surface value. The barrier layer was numerically calculated
by subtracting the mixed layer depth (MLD) from the isothermal layer. The isothermal
layer for the present study was defined as the depth at which the temperature decreased
by 1oC from its surface value.

The atmospheric data used was monthly mean climatology of the incoming short wave
radiation, wind speed, evaporation, precipitation and net heat flux on 1o x 1o grid, which
was obtained from National Oceanographic Centre (NOC), Southampton for the period
1980-1993. The remote sensing data used for the study was the chlorophyll pigment
concentrations from SeaWiFS during the period September 1997 to December 2007 and
the merged sea-level anomalies of Topex/Poseidon ERS1/2 series of satellites obtained
from AVISO live access server during the period October 1992 to January 2006. From
the sea-level height anomalies, velocities were computed assuming geostrophic balance.
The mixed layer and barrier layer variability were examined in the light of heat flux,
momentum flux (wind-stress curl) and fresh water flux (evaporation-precipitation) to
decipher the factors that are responsible for their changes.

The mixed layer depth (MLD) during the spring intermonsoon (March-April-May) was
the shallowest in the Bay of Bengal compared to the rest of the season. It varied between
10 and 25 m in March and April. This shallow MLD was driven by the strong
stratification induced by peak solar heating (280-290 W/m2) and subsequent highest net
heat gain by the ocean (150-160 W/m2). The low salinity waters (< 32.5 psu) in the
northern Bay (north of 18oN) during March-April made the upper ocean highly stratified.
The weak winds during this period were unable to drive deep wind-mixing due to strong
stratification and hence resulted in the formation of shallow mixed layer.

In the south, the comparatively deeper mixed layer (~ 35 m) seen west of 90oE was due to
the presence of high salinity waters (>34.5 psu) which made the water column less stable
and the moderate winds were able to initiate greater mixing leading to the observed deep
MLD. However, the deep MLD east of Sri Lanka was linked to the development of anticyclonic
circulation associated with the formation of subtropical gyre which begins in
May. This anti-cyclonic circulation drives down-welling and deepens the mixed layer.
The co-location of comparatively deep MLD (>25 m) and strong negative wind stress
curl (~ -20 x 10-8 Pascal/m) along the western boundary in April suggested the role of
wind stress curl in deepening the mixed layer. Note that the subtropical gyre was well
developed in April in the central and western Bay of Bengal, which also leads to downwelling
and augments the deepening of the mixed layer. The deep MLD in May in the
south, south of 4oN, was related to the downward Ekman pumping due to the negative
wind stress curl. In addition to this the time-longitude plot of sea-level height anomaly
along 4oN showed the propagation of Rossby waves during spring intermonsoon, which
also contributed in deepening the mixed layer.

During summer monsoon (June-July-August) though the wind speed was the highest in
the entire basin, the MLD was the shallowest in the northern Bay (~ 5m). An examination
of E-P showed that it was negative and the highest of all the season, implying excess
precipitation (in excess of 440 mm/month), in the northern Bay. In addition to the oceanic
precipitation, the influx of freshwaters from the rivers adjoing the Bay of Bengal also
contributed towards freshening of the surface waters of the Bay. The spreading of low
salinity waters (<32 psu) were seen from the northern Bay towards the south and east
with the progress of summer monsoon. The vertical profiles of stability parameter
showed that these low salinity waters strongly stratified the upper ocean. The warm upper
ocean with SST in excess of 28.5oC also contributed towards strengthening the
stratification. Hence, the winds though were the strongest of all the season, were unable
to break the stratification to initiate wind-driven mixing and deepen the mixed layer. The
shallow MLD seen around Sri Lanka was driven by the positive wind stress curl. The
positive wind stress curl was seen developing in May, which peaked in June and
collapsed by September. The upwelling associated with the positive wind stress curl
drives an upward Ekman pumping and this led to the observed shallow mixed layer
during summer monsoon. The band of deep mixed layer seen extending from the
southwestern region into the central Bay was linked to the advection of high salinity
waters from the Arabian Sea. This high salinity waters reduced the stratification of the
upper ocean as could be inferred from the stability parameter. Thus, the strong winds of
the summer monsoon combined with the less stratified upper ocean in the southern Bay
due to the intrusion of high salinity waters from the Arabian Sea were able to drive strong
wind-driven mixing. This was the mechanism which led to the formation of deep MLD in
the south. In addition to this, the high sea-level anomaly in the central and eastern part of
the southern Bay associated with the propagating Rossby waves also contributed towards
deep MLD.

As the summer monsson tapers off and the fall intermonsoon (September-October) sets
in, the shallow MLD which was confined to northern Bay, north of 18oN, was seen
extending southward to 15oN in October. This could be explained in the context of
changing atmospheric forcing from summer monsoon to fall intermonsoon. The short
wave radiation as well as net heat flux showed a secondary heating of the upper ocean
during fall intermonsoon and accordingly the SST was in excess of 29oC in October.
Though the E-P showed a rapidly decreasing precipitation, the surface salinity showed a
progressive decrease from that of summer monsoon and also a further southward
extension of the low salinity waters. This indicated that the shallow MLD in the northern
Bay and its further southward extension was linked to the presence of low salinity waters
and its advection southward. The winds over the Bay showed a drastic reduction in their
speed in the north during fall intermonsoon with the high wind speed confined to the
southern Bay. Thus, the deep MLD in the southern Bay was driven by a combination of
comparatively high wind speed and the presence of high salinity waters both of which
destabilized the water column.

The winter monsoon (November-February), in general, showed comparatively deep MLD
(~30-40 m) all over the Bay except in the north and eastern Bay. The shallow MLD (~5-
15 m) in the north and eastern Bay could be explained in the context of the presence of
low salinity waters (<32 psu) during November-December and associated strong
stratification. As the winter progressed, the E-P showed a net evaporation and with no
substantial input from rivers discharge the low salinity waters were confined to the
northern part during January-February. As a result the area of deeper mixed layer
expands further towards eastern boundary. The shallow MLD observed near the Sumatra
coast in January was driven by the strengthened positive wind stress curl and the
associated upward Ekman pumping. The deep MLD in the rest of the Bay was related to
the weakest stratification that occurred in the Bay during winter monsoon compared to all
other seasons. The wind speed, which showed a secondary peak in winter were able to
initiate deeper wind-mixing as the stratification of the water column was the weakest and
this gave rise to the deep mixed layer.

The seasonal variability of the barrier layer (BL) thickness showed strong coupling with
wind stress curl, freshwater flux and prevailing circulation of the basin. The nutrient
concentrations as well as chlorophyll a showed close correspondence with the changes in
the mixed layer. During spring intermonsoon the upper ocean was oligotropic and
showed very low chlorophyll a concentrations (~ 0.1 - 0.3 mg/m3). This was because of
the strongly stratified upper ocean due to the peak heating by incoming shortwave
radiation. The mixed layer was very shallow as the weak winds during spring
intermonsoon were unable to initiate strong wind-driven mixing. As a result there was no
vertical transport of nutrients to the surface layer from subsurface making the upper
ocean oligotrophic.

During summer monsoon the high nitrate as well as the chlorophyll a concentrations in
the Indo-Sri Lanka region was driven by the upwelling. The upward Ekman pumping
associated with upwelling transports sub-surface nutrients from the nitracline to the
surface layers and supports high biological productivity. In the northern Bay the higher
concentration of nitrate was supplied by the river runoff. The observed enhanced
chlorophyll a was supported by this nutrient input. Away from this region, the high
stratification of the upper layers due to the freshwater input from precipitation as well as
river runoff inhibited wind-driven mixing though the winds were strongest in summer
monsoon. Accordingly, the nutrient input from sub-surface to the upper ocean also was
curtailed. Thus, the shallow MLD and the oligotrophic upper ocean lead to the observed
low chlorophyll concentrations (~ 0.2 mg/m3) in the rest of the Bay. In fall intermonsoon,
the satellite-derived chlorophyll pigment concentrations showed pattern similar to that of
summer monsoon with a reduced concentration indicating the tapering effect of summer
monsoon. In winter, both the nitrate concentrations and the chlorophyll a concentrations
were high along the western boundary. The EICC which moved southward during winter
was capable of transporting some of the nutrients from the northern Bay towards the
south along the western boundary and could explain the nutrient enhancement which in
turn supported observed chlorophyll a concentrations.



Seasonal variability of the upper ocean driven by the atmospheric forcing and its regulation of nutrients and chlorophyll in the Bay of Bengal

AHMAD, SHOEB

 14 
ABSTRACT 
 
The solvent and structural influences on the kinetics of oxidation of aniline 
and para- (Me, OMe, COMe, NHCOMe, NO2, Br, Cl, F) and meta- (Me, OMe, 
COMe, Et, COOH, NO2) substituted anilines by two Cr(VI) oxidants viz. 
imidazolium fluorochromate (IFC) and nicotinium dichromate (NDC) have been 
carried out in neat organic solvents and in binary solvent mixtures. Electro-
oxidation of the chosen anilines was investigated in varying mole fractions of 
binary solvent mixtures with an aim to shed some light on the mechanism of the 
reaction. The kinetics of aromatic nucleophilic substitution reaction of these 
anilines with benzenesulphonyl chloride has also been carried out in neat organic 
solvents and in different binary solvent mixtures of varying compositions. The 
activation parameters have been computed following the reactions at different 
temperatures. The influence of structure and solvent on the reactivity has been 
analyzed using linear and multiple regression equations. The specific and non-
specific solvent-solvent-solute interactions on the reactant/transition state have 
been separated and quantified using the technique of correlation analysis. Reasons 
for the difference in reactivity, in all the above reactions, with structural changes 
in the substrate are also discussed. On the basis of the results of systematic 
correlation analysis, plausible mechanisms have been proposed for these reactions. 
The results of this study have been published in reputed journals.  
 
 


Structural studies on thermostable variants of Bacillus subtilis lipase evolved by in vitro methods

RAO, SUBBA

Abstract
	The study throws light on the production of betalaines from hairy roots of        B. vulgaris and biotransformations using C. frutescens normal roots for the production of vanillin related metabolites. Various aspects viz. polyamines, MJ and down stream process methodologies have been studied to enhance the metabolite production.

	It may be concluded from the present study that hairy roots can be used as a potential system for the production of betalaines. The organized nature and stable metabolite production makes them more amenable for bioreactor cultivation and scale-up studies. Among all bioreactors studied, acoustic mist bioreactor performed well with high growth and metabolite production. There was no mass transfer limitation observed in the acoustic mist bioreactor unlike others such as nutrient sprinkle and bubble column reactors. Osmolarity measurements proved reliable than conductivity for indirect estimation of biomass when the hairy roots were cultivated in bioreactor systems. 

	The studies on product enhancement (polyamines and MJ) and on-line extraction of pigment (using CTAB) provided an insight for integrating unit operations and developing a process for continuous operation and higher production of phytochemicals in bioreactor systems. In the present study it was found that the roots were still viable and showed normal growth even after CTAB treatment (0.002%) which would be an important observation to make this process a continuous one. 

	This study has shown the production of DOPA and dopamine, which are very important pharmaceutical compounds from hairy roots of B. vulgaris. Further studies with tyrosine feeding led to higher production of DOPA and dopamine. This study is of importance in understanding the biosynthetic pathway of betalaines derived from DOPA and dopamine. Use of oxygenase inhibitors such as caffeic acid and catechin formed the basis to envisage the enzymatic pathways in betalaine production. The higher production of DOPA and dopamine under the influence of caffeic acid and catechin might be because of the inhibitory action of these chemicals on tyrosinase preventing further conversions.

Future work would be oriented to the studies on metabolic block of enzymes such as DOPA dioxygenase to produce more DOPA, which is an important pharmaceutical compound.  Antisense RNA methodology for the enzyme DOPA dioxygenase would also be helpful probably leading the higher production of DOPA and dopamine.

The studies on biotransformation of phenylpropanoid compounds to various vanillin related metabolites using C. frutescens normal roots led to understand the biosynthetic pathways involving various chemical reactions. 

The higher biotransformation rates in cultures fed with ferulic acid complexed with BCD would be because of the increased solubility and availability of precursor. The formation of vanillin from ferulic acid might be via feruloyl-CoA through a reaction similar to -oxidation of fatty acids. Vanillic acid was formed by further oxidation of vanillin, whereas reduction yielded vanillyl alcohol. In the present study, biotransformation resulted in higher concentrations of vanilla flavour compounds in the medium than in roots. Thus, it would be appropriate to use a bioreactor system for down stream processing of the biotransformed products. The formation of vanillin from protocatechuic aldehyde involved O-methylation reaction and oxidation to produce vanillin and protocatechuic acid respectively.

It was observed that ferulic acid formation was more in the early stages after caffeic acid feeding as reflected by O-methylation reaction. When MJ was fed along with caffeic acid, it probably enhanced the CAOMT activity leading to increased biotransformed product. Further, ferulic acid upon oxidation probably led to vanillin production along with other metabolites. Veratraldehyde fed cultures showed maximum vanillin accumulation than other precursors. Veratraldehyde was demethylated to vanillin and parahydroxy benzoic acid. 
There was no capsaicin accumulation recorded in the studies indicating the non-existence of capsaicinoid synthase that is considered as a key enzyme in the synthesis of capsaicin.  The efficiencies of biotransformation in all the biotransformation studies ranged from 2-9% suggesting the possible channeling of precursors resulting in the accumulation of other compounds such as lignin.

Further work would involve the use of inhibitors of lignin biosynthetic pathway to divert the phenylpropanoid precursors to vanillin related metabolite pathways.
























FERMENTATION KINETICS OF PROTEASE PRODUCTION BY MICROBIAL SP.

Shamsan, Ebtisam Faisal S

Abstract
Platelets are the tiny corpuscular cells found in the blood along with erythrocytes and leukocytes.  They are continuously surveying the inner layers of intact blood vessels and play an important role in haemostasis.  Whenever there is any break / damage in the blood vessels, platelets are exposed to the damaged tissue resulting in a cascade of events like shape change, secretion and aggregation leading to the formation of a precise haemostatic plug.  But if these synergistic interactions take place in vivo, it results in the blockage of blood supply to essential organs causing cardio-vascular or cerebro-vascular complications.  It is reported that patients with atherosclerosis possess increased platelet activity.
Platelets are activated by a variety of agonists like ADP, epinephrine, collagen, thrombin, arachidonic acid, calcium ionophore A 23187 and ristocetin resulting in the shape change from discoidal to spherical.  Also platelets possess a unique property of ‘stickiness’, which is seen once they are activated resulting in the adhesion of platelets to one another.   
Platelet aggregation is a membrane-associated event and the involvement of platelet membranes is crucial due to their interaction with the various agonists.  A variety of dietary components influence the membrane characteristics like fluidity, stability and susceptibility to oxidative damage.  Membrane lipids are highly susceptible to peroxidation resulting in the damage to the membrane structure and function.  Spices the known food adjuncts possess anti-diabetic, anti-inflammatory, hypolipidemic and anti - lithogenic effect as per Ayurveda, the indigenous system of Indian medicine.  
Reports indicate that aqueous extracts of onion and garlic inhibited platelet aggregation by acting on the arachidonic acid metabolism.  Similarly, aqueous extract of ginger inhibited ADP, epinephrine, collagen and arachidonic acid induced platelet aggregation by reducing the formation of thromboxanes.
A dietary survey indicated that red chillies, turmeric, garlic, coriander and cumin are the most commonly used spices in Indian households.  Hence two of these – coriander and cumin were selected for the present investigation, as no information is available on the effect of these spices at present.   In principle the effect of these spices on human platelet aggregation, its membrane structure and function were investigated in vitro.  Experiments were carried out in vitro using platelets isolated from human blood.  
The results of this investigation are presented in three chapters.
Chapter 1:
	This chapter presents a brief review of existing literature on platelets, its formation, role in haemostasis and blood coagulation, platelet aggregation, lipid peroxidation, membrane bound enzymes like Na+K+ATPase and Ca+2Mg+2ATPase, membrane fluidity, fluorescence anisotropy, probes of membrane structure, anti platelet aggregating agents, abnormalities of platelet functions.
	A brief history of role of spices and their pharmacological role along with that of coriander and cumin is also presented.
Chapter 2:
	In this chapter the experimental methods employed in the present investigation is detailed.  


Chapter 3:
	The results of the present investigation haven been discussed in this chapter under five sub headings, which are as follows:
1.	Effect of coriander and cumin extracts on human platelets.
2.	Effect of coriander and cumin extracts on human platelet membrane lipid peroxidation and membrane fluidity.
3.	Effect of the coriander and cumin extracts on the kinetics of human platelet membrane bound enzymes.
4.	Sephadex fractionation of cumin and coriander extracts.
5.	Effect of other spices on human platelets.
1.	Effect of coriander and cumin extracts on human platelets:
 (a) Aqueous extracts of coriander and cumin were tested on human platelets for their inhibitory effect at different concentrations with a variety of agonists like ADP, epinephrine, collagen, calcium ionophore A 23187 and ristocetin to obtain the IC50 at one-minute incubation.
(b) The effect of increase in the duration of incubation from one minute to 2, 4 and 8 min were determined at the IC50.
(c) After aggregation the products like malondialdehyde (MDA) and serotonin released were assayed for all the spice extracts.
 (i) Raw extracts of coriander and cumin were tested on human platelets for their inhibitory effect and steps 1a – 1c were carried out.
(ii) Heat treatment of coriander and cumin.
The stability of the inhibitory activity of coriander and cumin were tested by subjecting them to heat treatment (boiling and roasting) as they may be susceptible to changes due to their chemical nature.  These extracts were used on human platelets and steps 1a – 1c were carried out.
(iii) Combination of coriander and cumin.
Spices are generally used in combinations.  In order to determine their synergistic effect, these two spices were taken in the ratio of 1:1. Raw and heat treated extracts were used on human platelets and steps 1a – 1c were carried out.
(iv) Processed coriander dhal extract was tested on human platelets and steps 1a – 1c were carried out.
(v) The pure isolated and selected components of coriander and cumin available commercially were tested on human platelets to assess their inhibitory effect and steps 1a – 1c were carried out.
(vi) Effect of extracts of coriander and cumin at IC50 were used to study their effect on washed platelets with agonists like ADP, collagen and A 23187.
2.	Effect of coriander and cumin extracts on human platelet membrane lipid peroxidation and membrane fluidity: 
Since lipid peroxidation adversely affects the biosystems, human platelet membrane lipid peroxidation was carried in vitro using iron-ascorbic acid system to assess the protective effect of extracts of coriander and cumin on lipid peroxidation.
Membrane fluidity using fluorescent probes was studied to understand the interaction of the extracts of coriander and cumin with human platelet membranes and its relationship to their inhibitory effect. 

3.	Effect of the coriander and cumin extracts on the kinetics of human platelet membrane bound enzymes:
As platelet aggregation is a membrane-associated phenomenon, studies were carried out to determine the effect of the raw and heat-treated coriander and cumin extracts on membrane bound enzymes Na+K+ATPase and Ca+2Mg+2ATPase.  Km and KI were determined for each of the extracts of coriander, cumin and their combination.
4.	Sephadex fractionation of cumin and coriander extracts:
The extracts of cumin and coriander were fractionated on sephadex G-50 and G-25, which yielded fractions with poor inhibitory activity.
5.	Effect of other spices on human platelets:
Initially other spices like cardamom, saffron, swallowroot, coriander leaf and curry leaf were screened for their inhibitory effect on human platelets.  Apart from that other reported spices like turmeric, ginger, garlic, cloves and cinnamon were also tested with agonists like ADP, epinephrine, collagen, A 23187 or ristocetin for comparison.




Ecobiology and Fisheries of an Economically Important Estuarine Fish,Sillago sihama (Forsskal)Ebtisam

Nagi, Hisham Mohamed

Abstract 

The thesis entitled “Synthetic studies on anthracyclinones, 1,4-anthraquinones and benzo[d]naphtho[1,2-b]pyran-6-ones” describes i) a direct synthetic access to 1,4-dihydroxyanthraquinones and exploratory studies towards anthracyclinones (e.g. 1) and a synthesis of two naturally occurring 9,10-anthraquinones ventinone A (2) and 4-methyl ether of erythroglaucin (also proposed structure of ventinone B) (3), ii) total synthesis of 9,10-dimethoxy-2-methyl-1,4-anthraquinone (4) and iii) model studies towards chrymutasins (5) and gilvocarcins (8). The key reactions investigated are Hauser annulation and carbonyl-ene reaction.



3-Phenylsulfanylphthalides (e.g. 9) readily react with p-benzoquinones (e.g. 10) in the presence of t-BuOLi in THF to furnish 1,4-dihydroxyanthraquinones (e.g. 11) in good yields and one-pot operations. This methodology has been applied to the total synthesis of naturally occurring anthraquinones, ventinone A (2) and 4-methyl ether of erythroglaucin (3).


Intramolecular carbonyl-ene reaction of 2-methallylated anthraquinone aldehyde 12, obtained in 10 steps has been effected for the synthesis of 13, core structure of idarubicin. Modest attempts have been made to extend the carbonyl-ene reaction, for the first time, to the synthesis of substituted naphthalene 16 and phenanthrene 17 as illustrated below. 



The first yet short synthesis of the unusual anthraquinone 4 has been achieved in 5 steps from p-benzoquinone using Kochi-Anderson radical methylation as a key step. During the course of the synthesis few unprecedented cyclopropaanthraquinones like 19, 20 and 21 have been synthesized and their chemistry studied. 




Cyanophthalide 22 has been shown to undergo a tandem annulation with methyl 5,5-dimethoxy-2-methyl-6-oxo-5,6-dihydronaphthalene 1-carboxylate (23), obtained in 10 steps from 6-methoxy-1-tetralone, to fabricate a chrymutasin chromophore 5 in one-pot operation and thus O-Methylhayumicinone (25) has been synthesized during this investigation. Both C and E rings are formed in one operation.

The tandem strategy has been applied to the regiospecific construction of the core structure of gilvocarcin chromophore (e.g. 26) from O-benzylcyanophthalide (27), prepared in 9 steps, and styryl sulfone 28, prepared in 6 steps. But, the total synthesis of defucogilvocarcin M (32) has been foiled, due to the failures in desulfonylation of 26.



Key words: Synthesis, anthracyclinone, 1,4-anthraquinone, cyclopropaanthraquinone, benzonaphthopyranone, Hauser annulation, intramolecular carbonyl-ene reaction, Kochi-Anderson radical alkylation reaction, Claisen rearrangement, Heck reaction.


3-Phenylsulfanylphthalides (e.g. 8a) readily react with p-benzoquinones in the presence of LiOtBu in THF to furnish 1,4-dihydroxyanthraquinones in good yields and one-pot operations.



A Hauser-initiated tandem annulation has been developed for the rapid regiospecific synthesis of benzonaphthopyranones. This strategy has been generalized with benzonaphthopyranones 26, 29, 32 & 35. It has also been employed in a short synthesis of chartarin (3) and O-methylhayumicinone (67).


The synthesis of 9,10-dimethoxy-2-methyl-1,4-anthraquinone, an unusual quinone, is achieved in 5 steps from p-benzoquinone. A Kochi-Anderson radical methylation features as a key step in the synthesis. The chemistry of a cyclopropa-1,4-anthracenedione is also described.



ENVIRONMENTAL STUDIES ON MANGROVE COVER CHANGES IN GOA, AND ITS RESIDENT CRASSOSTREA POPULATION

JAIN, SARIKA

  i 
INTRODUCTION 
Processing of food commodities to produce food products, which meet the 
safety and quality requirements of consumer, has been a challenge. Microbial 
hazards pose a significant threat to food safety, of which mycotoxin 
contamination is one of the main concerns of fungal infestation. Aflatoxin B1, 
one of the most potent mycotoxin was discovered in England in 1960 as a 
causative agent of aflatoxicosis in poultry. The discovery of aflatoxin led to 
screening and identification of more than 100 fungal metabolites, which are 
toxic to animals (Scudamore, 1994). Ochratoxins are such a group of toxic 
fungal metabolites, which has drawn much attention in the last few decades 
due to its implications in the nephropathy conditions in animals and humans in 
certain regions of the world. It is also studied for other toxic properties such as 
immunosuppression, teratogenecity and carcinogenicity. Ochratoxin A is 
classified by IARC as possible human carcinogen. The Joint FAO/WHO 
Expert Committee on Food Additives (JECFA) has recommended monitoring 
of this toxin and ochratoxigenic fungi in food commodities (WHO, 2002).   
The unprecedented growth of human population has necessitated the 
need for increasing the farm output and minimizing of the preharvest and post 
harvest losses of food grains. It is estimated that 5 - 10% of the food 
commodities grown in the world is rendered unfit for human consumption by 
fungal infestation (Pitt and Hocking, 1985). It is imperative to develop 
controlling strategies to reduce the pre and post harvest losses and microbial 
hazard in agricultural produce. Food industries are adopting the HACCP 
principles for producing quality food products free from hazards associated 
with food. Monitoring and control of the hazardous microorganism associated 
  ii 
with the food commodities are key factors for ensuring food safety. In this 
regard the present work is focused on developing methods for the 
identification of ochratoxigenic fungi and ochratoxins, prediction of the 
ochratoxigenic fungal behavior in foods, and control of ochratoxin A 
contamination.  
 
Objectives: 
 Evaluation of food commodities for ochratoxigenic fungal contamination 
and ochratoxin A contamination. 
 Identification and evaluation of ochratoxigenic potential of food isolates. 
 Immunological and DNA based methods for detection of ochratoxigenic 
fungi.  
 Immunological method for the determination of ochratoxin.  
 Microbiological studies on factors affecting ochratoxin A production. 
 Control of ochratoxin A contamination. 
  
The review of literature and research findings of the study undertaken to fulfill 
the objectives summarized here are presented in 8 chapters in the final thesis.  
 
 
 
 
 
 
 
  iii
Chapter 1.0 
Introduction 
Fungal spoilage of food commodities can occur both at the field and during 
storage. Fungal infestation is one of the important groups of spoilage 
organism responsible for pre and post harvest losses of agricultural produce. 
In this chapter consequences of fungal spoilage on loss of agricultural 
produce and their importance in food safety are emphasized. A list of food 
borne illness suspected to occur / caused by fungal spoilage is presented. 
The methods available for analysis of spoilage fungi and toxic metabolites are 
briefly explained. The chapter gives brief background information on threats 
and challenges posed by fungal spoilage. 
 
CHAPTER  2.0 
Review of literature 
The literature survey covering the current research status on ochratoxins and 
ochratoxigenic fungi is presented under the following major headings.  
 2.1  History and importance of ochratoxins  
 2.2  Ochratoxicosis  
 2.3  Ochratoxigenic fungi and their taxonomy  
 2.4  Chemistry of ochratoxins  
 2.5  Natural occurrence of ochratoxigenic fungi 
 2.6  Natural occurrence of ochratoxins  
 2.7  Biosynthesis of ochratoxins  
 2.8  Mycotoxigenic fungal study  
 2.9  Methods for determination of ochratoxins.  
  iv 
2.10  Factors affecting fungal growth and ochratoxin A production.  
2.11  Biological data on ochratoxin A  
2.12  Fate of ochratoxin A during food processing  
2.13  Degradation of Ochratoxin A  
2.14  Prevention of Ochratoxin A Contamination  
2.15  Risk assessment, Regulation and Legislation  
2.16  Microbiological Safety and Quality   
The literature review revealed scanty reports from Indian subcontinent 
on ochratoxin contamination in food commodities of which only few reports 
have detailed on ochratoxigenic fungi. No report is available on screening of 
food mycoflora other than A.ochraceus group for evaluation of ochratoxigenic 
potential from Indian subcontinent. The reports of identification of new species 
of ochratoxigenic fungi other than A.ochraceus group emphasize the need for 
evaluating food commodities for fungal isolates other than A.ochraceus group 
for ochratoxin elaboration.  
The literature review revealed lack of specific microbiological 
differentiation medium for ochratoxigenic fungi belonging to A.ochraceus 
group. The comparatively slow rate of growth of species belonging to 
A.ochraceus group of fungi also makes it difficult to identify these fungi in 
conventional general-purpose mycological medium. This indicated ample 
scope for development of rapid and specific detection method for these fungi 
for regular monitoring and to study the influence of biotic and abiotic factors 
on these fungi. The review also revealed ochratoxin A contamination in spices 
apart from the common major commodities such as maize, coffee and feed 
commodities. This emphasizes the need for simple, rapid screening and 
  v 
quantification methods for constant monitoring of various food commodities to 
ensure food safety. The biocontrol of the A.ochraceus group of fungi is limited 
to use of preservatives, plant extracts and irradiation. The data on 
degradation/ biotransformation of this toxin is limited to fate of this toxin during 
food processing environment and degradation by few microbial cultures.  
 
Chapter 3.0 
Prevalence of ochratoxigenic fungi and natural occurrence of 
ochratoxins in foods 
Ochratoxins are not only potent nephrotoxin but in recent years are also 
considered to be a carcinogen, which prompts the food analysts to monitor 
this toxin in food commodities. The investigation revealed that the distribution 
of ochratoxin producers is wide spread among the food commodities like 
cereals, spice, coffee beans and feed materials. This study show that 
A.ochraceus species is the dominant fungus producing ochratoxins along with 
other members of the A.ochraceus group that included A.sulphureus, 
A.auricomus, A.melleus and A.ostianus. The natural occurrence of ochratoxin 
A was limited to a sample each of poultry feed, groundnut cake and coffee 
bean among the 73 different food commoties tested representing cereals, oil 
seed, feed, beverage and spice (4.1%). This is well in agreement with other 
similar work reported in Indian sub continent, which was in the range of 1 - 
6%. However the incidence of A.ochraceus fungi was found in 20.5% of food 
commodities, in which 59.5% of the isolates belonging to A.ochraceus group 
had the potential to produce this toxin ranging from 0.4 ppm to 200 ppm. 
Distribution of ochratoxigenic fungi with high toxigenic potential in variety of 
  vi 
foods reaffirms the need for constant monitoring of this toxin in food 
commodities. 
 
Chapter 4.0 
Immunological methods for detection of ochratoxigenic fungi 
The main hurdle in fungal detection and identification is the need of 
mycological expertise and need of specific medium that can be used 
effectively to enumerate and identify the specific toxigenic fungus. The 
toxigenic fungus varies in their growth requirements such as water activity, 
temperature, pH etc and hence no single medium can be used for 
identification of all fungus. Moreover the fungus also differs in growth rate on 
the common enumeration media such as PDA, Czapak Dox agar, and Malt 
extract agar and hence the fast growing organisms such as A.niger and other 
fungus dominate and suppress the slow growing fungi. Two immunoassay 
protocols were optimized for detection of ochratoxigenic fungi, which is 
presented in chapter 4A and chapter 4B. 
 
Chapter 4A  
Microplate immunoassay for monitoring of A.ochraceus in 
foods 
Indirect non-competitive microplate immunoassay developed specifically to 
detect the A.ochraceus group of fungi in this study has direct application. 
Immunological direct detection of fungal biomass is possible and is 
advantageous over other chemical methods of indirect fungal biomass 
estimation such as ergosterol, glucose amine etc.  
  vii 
The sensitivity of the method was ≥ 0.2 µg fungal biomass per mg substrate 
and could be used to detect fungal spore at ≥ 4 log10 cfu/g. The spore 
enrichment technique was adopted to increase the detection limit for spores in 
the immunoassay, which enabled detection of spore at ≥ 3 log10 spores/g.  
The method was used for monitoring and comparison of A.ochraceus 
growth in different food commodities representing beverage, spice and feed 
commodities. poultry feed, which is a mixture of various feed ingredient 
supported highest fungal biomass and toxin production followed by coffee 
beans and chili in the substrates studied. The method was also applied to 
estimate fungal biomass in maize under the influence of temperature, 
moisture and inoculum in the predictive microbiology study of A.ochraceus 
behavior in maize (chapter 7).   
 
Chapter 4B 
Dot-binding immunoassay for detection of A.ochraceus in 
foods 
The non-competitive dot-binding ELISA developed for the detection of 
ochratoxigenic fungi has potential to be used as an alternative to microplate 
immunoassay in field condition. The immunoassay was sensitive to 
A.ochraceus biomass at ≥1 µg/ml in pure culture studies. The immunoassay 
can be used for detection of A.ochraceus infestation at ≥5 µg/mg substrate in 
chili and poultry feed whereas in coffee beans the detection is possible at ≥10 
µg/mg substrate. An indirect competitive dot-binding immunoassay was also 
optimized, which has a limit of detection at 0.2 µg A.ochraceus biomass per 
ml in pure culture studies.   
  viii
Chapter 5.0 
PCR based detection method for detection of ochratoxigenic 
fungi 
Molecular detection of ochratoxigenic fungi by PCR technique has been 
optimized. The sequence on the small subunit ribosomal RNA gene of 
common fungal genera were compared with that of A.ochraceus and two 
primer pairs were designed for specific amplification of 18S rRNA gene in 
A.ochraceus. The designed primer sets OT1 and OT2 targeted 906 bp and 
353 bp fragments respectively in the 18S rRNA gene. The PCR was 
optimized for specific amplification of A.ochraceus and related species 
belonging to A.ochraceus group. The method is specific to A.ochraceus group 
of fungi as substantiated in the study by experiments carried out with DNA 
extracted from common foodborne fungal cultures in pure and mixed culture, 
as well as directly in food commodities. The minimum quantity of biomass and 
spore concentration required for successful detection by the PCR was 
determined. The method was sensitive to A.ochraceus spore at ≥ 4log10 
spores/g and 10 mg biomass. Studies were conducted to increase the 
sensitivity of the assay for detection of A.ochraceus spore by enrichment 
technique. The A.ochraceus spores at concentration of 100 spores/g could be 
detected by the PCR following the spore enrichment prior to DNA isolation. 
  
 
 
 
 
  ix
Chapter 6.0 
Immunological method for determination of ochratoxin A 
Immunoassays for detection of ochratoxin A in food have been optimized. 
Ochratoxin A-BSA conjugate was used as an immunogen to raise antibodies 
in rabbit and as well as in hens. The sensitivity of antibody (IC50) for OTA was 
15ng/ml and 40ng/ml for antibodies elicited in rabbit and egg yolk respectively 
in the indirect competitive immunoassay. Protocols for ochratoxin estimations 
by ELISA were optimized using poultry feed as food substrate. Recovery 
studies on ochratoxin A (OTA) spiked poultry feed had a recovery at 66 -
125% of added OTA with a detection limit of 5ppb in the ELISA. Market 
samples of poultry feed samples were analyzed by ELISA and HPLC. ELISA 
estimations compared well with the HPLC method in all the samples analyzed 
without any false positive or negative results. The results with hen egg yolk 
antibody indicate the potential of using hen egg as a source for large-scale 
production of antibodies against mycotoxins.  
 
Chapter  7.0 
A predictive microbiological study – Interrelationships of 
temperature, moisture, and inoculum level on behavior of 
toxigenic A. ochraceus 
Influence of moisture, temperature, and inoculum on A.ochraceus growth and 
ochratoxin A elaboration in maize was studied using three factorial Central 
Rotatory Composite Design. Multiple linear regression analysis of fungal 
responses under the influence of different combination of statistically 
designed variables was carried out and polynomial regression equation was 
  x
derived to predict the fungal responses at different variables. The study 
revealed that the fungal biomass correlates more positively with ochratoxin 
elaboration than with viable count of fungi. The coefficient of correlation 
between biomass-viable count, biomass-ochratoxin A elaboration and viable 
count-ochratoxin A elaboration were 0.79, 0.79 and 0.57 respectively. The 
bias factor of 1.0 and 0.97 for predictions for fungal growth and ochratoxin A 
elaboration study respectively indicate that the predictions obtained within the 
experimental domain are dependable. The predictive model when used with 
the immunological method for determination of fungal biomass has potential 
for application in food processing industries to predict the behavior of 
ochratoxigenic fungi in specific food environment.  
 
Chapter 8.0 
Control of ochratoxigenic fungi and degradation of 
ochratoxins by microbiological method 
The work relating to screening of organism, which are antagonistic towards 
ochratoxigenic fungi and organism that can degrade ochratoxin has been 
presented in chapter 8A and chapter 8B respectively.  
 
Chapter 8A  
Identification, Isolation and application of antifungal 
metabolite 
Bacillus species, identified as Bacillus badius, which was antagonistic against 
fungi belonging to A.ochraceus group and few other common fungi, was 
isolated. The organism produced an extracellular metabolite, which had a 
  xi
molecular weight between 2000–10000 Daltons, was found to be heat stable 
to temperature up to 80oC and was also found to be active in a wide pH range 
of 2.5 - 10. The metabolite was resistant to the action of enzymes viz., trypsin, 
and lysozyme. The metabolite exhibited inhibitory activity against A.ochraceus 
growth in broth culture as well in solid food substrate. The metabolite inhibited 
85% of A.ochraceus biomass and 100% inhibition of ochratoxin A elaboration 
in maize compared to a control experiment without the metabolite. 
 
Chapter 8B  
Degradation of ochratoxin A by Pseudomonas species 
About 10 Pseudomonas species were screened to evaluate their potential for 
degradation of ochratoxin A. Cell free extracts from two of the Pseudomonas 
fluroscens strains screened exhibited potential to degrade ochratoxin A. The 
temperature and time for degradation of ochratoxin A was optimized and 
degradation of ochratoxin A at different concentration under optimum 
conditions was evaluated. The cell free extract exhibited potential to degrade 
80 -100% of added ochratoxin A at tested concentrations ranging from 0.2 to 
500 µg. A bioassay with Bacillus subtilis, which was sensitive to ochratoxin A 
was used as an indicator organism to study the toxicity of the degraded 
product of ochratoxin A. The microbial assay revealed that the OTA degraded 
byproducts is not toxic. 


DERIVATION OF SOUND SPEED PROFILES USING IN SITU AND REMOTE SENSING DATA

Saafani, Mohammed Ali Yahya Al

Abstract of the thesis entitled “Synthesis, characterization and olefin polymerization reactivity of some early and late transition metal complexes” 

This thesis describes the synthesis and characterization of selected early and late transition metal complexes and their usefulness as catalysts for the polymerization of ethylene.

Polymerization of ethylene was studied using copper(II) complexes based on -diimine, bis(oxazoline) and bis(benzimidazole) ligands. (-diimine) copper(II) complexes bearing an acenapthene ligand framework exhibited moderate activity for polymerization of ethylene when activated with MAO at 5 bar ethylene pressure and maximum activity was observed for the complex with ortho-isopropyl substituents.  The complex bearing an aliphatic -diimine ligand framework was inactive for polymerization of ethylene even at 5 bar pressure indicating that nature of the chelating ligand plays an important role in the generation and stabilization of the active species.  Poly(ethylene)s produced by the copper complexes were linear and exhibited high molecular weights and narrow molecular weight distributions which implies the single site nature of the active species. The active species is proposed to be a copper(II) centre with -Cl or -Me bridges.  Bis(oxazoline) and bis(benzimidazole) copper(II) complexes were not active for polymerization even at 5 bar pressure.

Group 4 (Ti, Zr, Hf) metal complexes of trans-2-[9-(H) fluorenyl] cyclohexanol were studied for the polymerization of ethylene with MAO as cocatalyst.  They exhibited moderate catalyst activity for polymerization of ethylene and activity decreased in the order Zr>Ti>Hf.  These complexes exhibited activity even at 100C.  The zirconium complex produced poly(ethylene)s with exclusively saturated chain ends when the polymerization was conducted at 25-60C.  The saturated chain ends may result from chain transfer to aluminium.  However, at temperatures above 80C, a mixture of solid polymer and soluble oligomers was obtained.   The solid polymer consisted of long chain branches that are proposed to be formed by -H transfer followed by reincorporation of the resulting macromonomers into the growing polymer chains.  When the polymerization was conducted at 100C, methyl branches were also observed in the polymer in addition to long chain branches.  These methyl branches may result from -H transfer followed by 2,1-insertion of the -olefin into the [Zr-H] species. Both saturated chain ends and vinyl unsaturation were observed in poly(ethylene)s produced at higher temperatures that implies that -H transfer becomes more competitive with chain transfer to aluminium.  The soluble oligomers produced at higher temperatures were found to consist of a mixture of linear -olefins and long chain alkanes.  

Half sandwich fluorenyl amido complexes of titanium and zirconium based on ligands bearing aromatic substituents on the amido nitrogen were synthesized and employed as catalysts for the polymerization of ethylene with MAO as the activator.  The complexes exhibited moderate catalyst activity and resulted in linear, high molecular weight poly(ethylene)s.  Catalyst activities of these complexes were lower when compared to similar complexes bearing alkyl substituents on the amido nitrogen. It is speculated that resonance effect of the aromatic substituents may be responsible for the lower catalyst activities.



A.Rajesh								Dr.S.Sivaram
(Research scholar)							(Research supervisor)

Date:



Physical Oceanography of the Gulf of Aden

Bafana, Amit

Abstract

Microbial Degradation and Detoxification of Synthetic Dyes Commonly Encountered in Indian Environment

Azo dyes are the largest group of colourants constituting 70% of all organic dyes produced in the world. Around 2-50% of these dyes are lost in the effluent during dyeing process. Since they are designed to resist fading by different physical, chemical and biological agents, they are not degraded by conventional treatment processes and enter the environment. These coloured discharges are not only aesthetically unpleasing and drastically affect photosynthesis in aquatic ecosystem, but also pose ecological and public health risk. Moreover, many of the azo dyes and/or aromatic amines produced by their degradation are potent carcinogens, and are bioaccumulated and bioconcentrated along the food chain. This has induced considerable research in the use of biological systems for treatment of dye wastewater.
	Due to their wide usage, persistence and toxicity, azo dyes were selected as the subject for this study. The study mainly focuses on biodegradation and detoxification of two benzidine-based azo dyes: Direct Black 38 (DB38) and Direct Red 28 (DR28). Four other azo dyes, which are readily available and commonly used in India, were also chosen to check the effect of dye structure on degradation by the microorganisms isolated in this study. These were Direct Blue 1 (DB1), Direct Blue 6 (DB6), Direct Blue 15 (DB15) and Acid Yellow 36 (AY36).
The objectives of this study were:
	Isolation of dye decolourizing bacteria and characterization of the basis of their dye decolourizing activity.
	Identification of the intermediates formed by dye decolourization and comparison of their toxicity with that of dye to confirm detoxification.
	Determination of suitability of an industrial dye wastewater treatment plant by characterization of biodiversity of its dye decolourizing microbial population.
	Cloning and sequencing of azoreductase genes responsible for dye decolourization.
	Determination of the sequence variation and phylogenetic relationship among various azoreductase genes available in the database.




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