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Item Investigation of the effectiveness of mother enzyme on model organic waste material by enzyme degradation(University of Sri Jayawardenepura: USJ(Main)., 2002) WIMALADHARMA, AGIIIt is reported that at least 22 human diseases are associated with solid wastes. (Arne, Jeffery \& Arber 1982). Therefore continuous disposal of solid waste and their collection creates a severe environmental health hazard. As such, it is of utmost importance to avoid this problem before it is too late to do anything about it. In Sri Lanka the major percentage of municipal solid waste consist of organic waste. Hence one way of overcoming this problem is to increase the rate of degradation of organic material in which enzyme and micro organisms play a big role. Mother Enzyme (ME) is said to be one such mixture of enzymes and micro organisms. Objectives of this research was to identify the constituents of the ME, to determine the effectiveness of the ME on degradation of different types of organic wastes, and to determine the effective levels of ME for different types of organic wastes. Laboratory experiments were done at the Department of Forestry and Environmental Science, Department of Microbiology of Faculty of Medicine and the Food Science laboratory of the Department of Chemistry of the University of Sri Jayawardenepura. Results were analyzed using Mann-Whitney U-Test and Wilcoxon's Signed Rank Test. Macconkey agar, Nutrient agar, and Sabouraud Dextrose Agar were used to find the total number and the types of micro-oranisms present in the ME. Effect of Mother enzyme on different types of organic waste were done by using Farm (cattle and poultry) waste, Kitchen wastes and Fruit industry wastes in the presence and in the absence of ME with replication. Weight reduction with time was noted down. To estimate the effect of ME on model organic waste, wheat flour, Egg Yolk and coconut oil were used. Alanine, Glucose and Acid value estimation were done to estimate the rate of degradation of the above macro molecules in the presence of and in the absence of ME. In order to find out the effective levels of ME Farm (cattle and poultry), Kitchen wastes and Fruit industry wastes were used with replication. 2g/1kg and 0.5g/1kg of mother Enzyme was given and weight reduction was noted down. From the Microbiological analysis, Staphylococcus aureus, Escherichia coli and Mucor, were found to be present in ME and the total number of cells present was 13x101 g-1. Since it was unexpected to find E.coli in Mother Enzyme we can conclude that E.coli. Has come into the sample by some kind of contamination that has occurred either in the original sample of Mother Enzyme we used as the test material or the during laboratory testing. Presence of Mucor, common contaminant fungi is also surprising. From the study it was found that the break down of model organic matter-fats and proteins occur quite fast in the presence of mother enzyme. But breaking down of carbohydrates is not accelerated significantly by mother enzymes. Increase or decrease levels of ME have no significant effect on mixed organic waste due to our consumption pattern most of organic waste generated in Sri Lanka are of carbohydrates origin. This may be the reason why there is no significant effect due to ME on different organic waste found in Sri LankaItem Ayurvedic Drugs(University of Sri Jayawardenepura: USJ (Main)., 2002) TENNAKOON, MSGAyurveda plays a significant role in the health care in Sri Lanka. Within the social context in which Ayrveda is practiced in the present day, the standardization and quality assurance of Ayurvedic drugs is urgent and imperative.Dasamoolarista (DMA) is a complex drug containing over 60 ingredients. Test for identify in complex herbal drugs such as DMA can be devised through chromatographic methods to identify specific marker compounds which can be correlated with specific plant ingredients and representative compounds for groups of plants ingredients. A strategy for identifying such marker compounds by comparing the thin layer chromatograms of drugs (Arista) having closely related formulae, was developed. Thin layer chromatographic systems to detected the following plant ingredients through the marker compounds and representative compounds shown in parenthesis were developed; Aegle marmelos Correa. (Umbelliferone); Plumbago indica L. (Isoshinanolone and Epiisohinanolone); Saussurea lappa C.B. Clarke. (Dehydrocostus lactone and Dihydrodehydrocostus lactone); Glycyrrhiza glabra L. , Pterocarpus marsupium Roxb., and Myroxylum balsamum (L.) Harms. (Isoliquiritigenin); Rubia cordifolia L. (Alizarin and Purpurin); Eugenia caryophyllata Thumb, and Cinnamomum verun Presl (Eugenol);Woodfordia fruticosa (Linn.) Kurz., Vitis vinifera L. and bees honey (Quercetin and Kaempferol). It is proposed that tests for strength (potency) of drugs such as DMA of unknown pharmacological action, can consist of quantitative measurements of compounds of high biological activity found in the drug and specific marker compounds irrespective of their biological activity. Analytical methods based on TLC densitometry, HPLC and GLC were used to quantify gallic acid, isoliquiritigenin, umbelliferone, dehydrocostus lactone, dhydrodehydrocostus lactone, 5-hydroxymethylfurfral, isoshinanolone and epiisoshinanolone in DMA. The analytical methods were shown to be precise and accurate. These methods were then used to study the variability in composition of different commercial brands of DMA, and of different manufacturing batches of DMA of the same brand. Inter batch and inter brand variability was high, indicating a significant variation in the quality of crude drugs used in the manufacture of the drug. Changes in the level of dehydrocostus lactone, gallic acid and 5-hydroxymethylfurfural during the different stages of manufacture were monitored. It was concluded that levels of gallic acid and 5-hydroxymethylfurfural were more suitable as parameters for process control than those of dhydrocostus lactoen.Item Ayurvedic preparations containing cannabis some analytical aspects of pharmacological and legal importance(University of Sri Jayawardenepura: USJ(Main)., 2000) ABEYSINGHE, DMUJAyurvedic preparations containing varying amounts of Cannabis are freely available in Sri Lanka. These narcotic preparations are being abused by youngsters including school children which poses a threat not only to their health but also to society. However current narcotics legislation in Sri Lanka does not cover these preparations. "Madana Modakaya" (MM) is the most popular preparation currently available.The aim of this study was to develop a method to quantify the ?9- Tetrahydrocannabinol (?9-THC) levels in MM , to use this method to analyse market samples and to study how the manufacturing process affected the final level of ?9-THC in the product. It is hoped that this data will enable the authorities to legislate for permissible levels of ?9- THC in MM, and that the method developed would be used as the official method of analysis. A method to quantify ?9-THC in MM was developed. The method which involved solvent extraction and column chromatography followed by GC, was validated by the standard addition recovery method. In the study of the methods for the preparation of MM, TLC densitometry was used in quantifying ?9- THC, cannabidiol and cannabinol. A laboratory sample prepared according to the ayurvedic pharmacopoeia was used as the standard preparation. A field survey revealed that there were over 20 brands of MM in the market. The price and weight of a packet (single dose) varied from 3-5 rupees and 5.70 - 10.01 g respectively. The ?9- the content varied from 0.0183 percent to 0.0747 percent (w/w). None of the samples of the 23 brands studied contained atropine, indicating that the popular belief that MM is adulterated with Datura seeds is not true. The Ayurvedic pharmacopoeia describes two methods for "detoxification" of Cannabis prior to incorporation in the drug. It was shown that one of the method, namely frying in ghee, resulted in the reduction of ?9-THC approximately by 50 percent. Based on data obtained on the standard sample and data on the highest levels of ?9-THC found in local Cannabis, a permissible maximum of 0.031 percent (w/w) of ?9-THC in MM is proposed.Item Investigation of the effects of a traditional Sri Lankan Medicine on Hepatocarcinogenesis(University of Sri Jayawardenepura: USJ(MED)., 2003) IDDAMALDENIYA, SSHepatocellular carcinoma is among the eight leading causes of cancer deaths worldwide with a clear tendency to increase further. Therapeutic possibilities for this are very limited and prognosis is usually poor. Several plant-based treatments are being recommended for cancer patients by traditional medical practitioners of Sri Lanka. However, none of these has been subjected to scientifically controlled investigation to validate their anti-cancer potential. One of these is a decoction comprised of Nigella sativa seeds. Hemidesmus indicus root and Smilax glabra rhizome. In the present investigation protection against Diethylnitrosamine (DEN) - induced hepatocacinogenesis was investigated in Wistar rats by the decoction using the medium term bioassay of Ito, based on a two-step model of hepatocarcinogenesis. In previous studies, garlic has been shown to protect, rat liver against DEN-induced carcinogenesis. Therefore, it was used as a positive control in the present study. Objectives of the investigation were to determine:- 1. whether a decoction of Nigella sativa, Hemidesmus indicus and Smilax glabra ubhibits glutathione S-transferase (GST-P) expression in rat liver. 2. Whether this decoction has any anti-tumour potential. 3. Whether this decoction has any toxic side effects. 4. The mechanisms/s of action/s by which the decoction mediates its anti-tumour activity. Four studies (Studies 1,2,3 and 4) were conducted to achieve the above four objectives. In study 1, the short-term effects of the decoction on GST-P+ expression in rat hepatocytes were investigated. Carcinogenic potential in study 1 was scored by comparing the number, area and staining intensity of GST-P positive foci and number of cells/cm2 of the foci in the livers of rats treated with the decoction (test 1 and test 2) or garlic (positive control, control 2) for 10 weeks with those of the corresponding group (control 1 ) of rats given DEN and distilled water. Decoction dose 1(4g/kg body weight/day) corresponding to the normal therapeutic dose, was administered to the test group of rats, while dose 2, was given to the test 2 group of rats, provided a higher dose (6g/kg body weight/day). Treatment with decoction 1 reduced significantly, (a) the number and area of GST-P positive foci, (b) number of cells/cm2 of foci, (c) staining intensity of GST-P positive foci (p\<0.01) compared with animals in control 1. Treatment with decoction dose 2 resulted in further significant reduction in the above parameters (P\<0.001). The reduction mediated by dose 2 was similar to that produced by garlic (20mg/kg body weight/day). In study 2, the effects of long-term treatment (for 9 months) of the decoction on tumour development were investigated. Two groups were used-DEN only treated group (DEN-control) and DEN+ decpction treated group (6g/kg body weight/day; test study). During the post mortem of rats after nine monts, one hepatocellular adenoma (HA) was found in the DEN control group. Haematoxylin and Eosin staining of liver sections confirmed the HA and revealed altered hepatocyte nodules which may progress to HA.DEN and decoction treated group showed no HA. Reticulin stain was done to confirm the HA and to see whether it has progressed up to the hepatocellular carcinoma (HCC). DEN control group showed well-preserved reticulin framework of normal liver, which confirms that HA , has not yet progressed up to HCC. Test groups showed expected normal liver recticular pattern. The effects of the decoction were investigated in study 3. Treatment with decoction dose 1 or dose 2 for three months had no adverse effects on the liver function (as assessed by its effects on serum levels of alanine and anspartate aminotrasnferase and alkaline phosphatase). Histopathological studies indicated that no significant histological changes had occurred in any of the major body organs (liver, kidney, lung heart, stomach and duodenum) investigated. No significant changes in haematological parameters (red blood cell count, white blood cell count, haemoglobin concentration, packed cell volume, mean corpuscular haemoglobin, mean corpuscular volume and mean corpuscular haemoglobin concentration) were observed during treatment with this decoction. The investigations also demonstrated that the decoction did not possess anti-ovulatory, anti-implantation, and anti-spermatocytogenic properties. The LD50 study showed that event at a dose of 40 times the highest dose (6g/kg body weight/day), used in other experiments of the study, the decoction did not cause any mortality. Long-term (3 months) treatments of the study, the decoction did not cause any mortality. Long-term (3 months) treatment with the decoction did not cause any changes in average feed consumption, average body weight gain, and body weight: liver weight ratios and the general behaviour of the animals. In study 4, preliminary investigations were carried out to determine possible mechanism of action by which the decoction mediates its anti-cancer effects. Tests done for anti-oxidant activity showed that the decoction significantly increases the activity of blood glutathione peroxidase and superoxide dismutase (p\<0.05) with little radical scavenging activity. Studies on immunomodilatory activity indicated that the decoction could stimulate production of T lymphocytes and NK cells (CD8 and NK receptors) although the results were not statistically significant. Overall results indicate that the decoction comprised of N.sativa seeds, H.indicus root and S glabra rhizome can protect against chemically induced hepatocarcinogenesis with no significant toxic effects even when it was administered for a period of three months. Antioxidant activity and immunomodulation are two possible mechanisms by which the decoction mediates its anti-carcinogenic activity.Item Effects of garlic (Allium sativum) on chemically induced hepatocarcinogenesis in normal and hypercholesterolaemic Wistar rats(University of Sri Jayawardenepura: USJ(MED)., 2000) SAMARANAYAKE, MDPHepatocellular carcinoma is one of the most common cancers in the modern world and it is prevalent in Asia. Environmental and dietary factors play an major role in the development of many cancers. Dietary modification can also play important role in reducing the risk of cancer. Garlic (Allium sativum) is a well known medicinal herb and a food item that has been used all over the world since pre historic times. Previous studies using cancer cell lines and animal models have indicated that garlic and some of its sulfur compounds are potential anticarcinogens. The present study was carried out to investigate the effects of a therapeutic dose (20mg/kg body wt./day) of garlic on chemically induced hepatocarcinogenesis in normal and hypercholesterolaemic Wistar rats. In the fist phase of this study the inhibitory effects of a therapeutic dose (20mg/kg body wt./day) of garlic on diethylnitrosamine (DEN)- induced neoplasia of the liver were examined in normal male Wistar rats. Medium-term Bio assay system of Ito based on the two step model of hepatocarcinogenesis was used as the assay method. In phase II, the effects of garlic on hepatocarcinogenesis in hypercholesterolaemic Wistar rats were investigated. The selection of hypercholesterolaemic model was based on the available evidence for the relationship between dietary fat and the development of cancer. Hypercholesterolaemic model was developed by feeding rats with a 0.5 percentage cholesterol-enriched diet for a period of two weeks. Rats having serum cholesterol level \>120 mg/dl. Were considered as hypercholesterolaemic (normal range 75ñ10 mg/dl) and the cholesterol-enriched diet was continued throughout the experiment. Carcinogenic potential was scored by comparing the number and area of induced Glutathione S-transferase placental form positive (GST-P+) liver foci as well as histopathological examination of liver sections. Daily treatment with garlic markedly reduced the number and area of GST-P+ foci (48 percent inhibition and 49percent respectively) as compared with the control group of animals receiving distilled water. Significant inhibition of induction of GST-P+ foci (34 percent inhibition in number and 44 percent inhibition in area) due to garlic treatment was also observed in the hypercholesterolaemic group of rats. However percentage inhibition was higher in garlic treated normal rats than garlic treated hypercholesterolaemic rats. Also it was evident from this study that hypercholesterolaemic rats are more susceptible to induction of liver GST-P+ foci. Results of GST-P+ expression were supplemented by histopathological examination of liver sections of garlic treated normal as well as hupercholesterolaemic Wistar rats. Granular and vacuolar degeneration were used as parameters in assessing histopathological alterations. In normal rats, garlic treatment reduced the pathological alterations in liver sections caused by DEN. Similar results were obtained from experiments with hypercholesterolaemic rats. Hence the results of this study provide strong supportive evidence for the anticarcinogenic activity of garlic.Item The Diversity, bioactivity and structural studies of flabelliferins from (Borassus flabellifer L.) fruit pulp(University of Sri Jayawardenepura: USJ(MED)., 2002) ARIYASENA, DDPalmyrah fruit pulp (PFP) has a total potential production of 15-20 k tones. Annum-1. However, the bulk of PFP goes to a waste on account of : (1) inadequate basic knowledge in processing and (2) the presence of a bitter principle and bioactive factors now collectively known as flabelliferins. These are steroidal saponins. The overall objective of this study was to collect chemical data of flabelliferins from PFP to promote its wider utilization. There are two possible major avenues for utilization: (1) debittering the fruit pulp to give jams, cordials etc., (2) fermenting the fruit pulp to obtain potable alcohol. The bitter flabelliferin, F-II, a tetraglycoside is an inhibitor to the Na+/K+ pump. The triglycoside FB is a microbial inhibitor (yeast and selected bacteria). The triglycoside Fc and the diglycoside FD are inactive flabelliferins. F-I was reported to be a tetraglucoside. In order to support the utilization of PFP , it was important to study the flabelliferin profiles of varying fruit pulps and attempt to correlate them to an easily distinguishable feature of the fruit or the fruit pulp so as to aid in the selection of the best mode of processing for utilization. The study showed that flabelliferin profiles of specimens collected varied considerably in morphology. There was a common type, type -I (size, medium to big; colour, black; pericarp, rough with brown longitudinal striations; distal side, black) and three other types, type - II, typeIII and type IV. There was no correlation between falbelliferin profile and morphological type, colour of fruit pulp, carotenoid content (total absorbance) and the carotenoid present (?max). As expected bitter fruit pulps contained large amount of F-II. There was great diversity in flabelliferin profile ranging from 2 flabelliferins (for example, Chilaw) to more than 10 flabelliferins ( for example, Polonnaruwa). There appeared to be a correlation with location but this line of study was not pursued. The diversity of flabelliferin profiles made previously documented procedures of separation not universally applicable to all PFP's. When the flabelliferin mixture is complex (for example, the Polonnaruwa sample containing at least 10 flabelliferins) a better separation technique was needed. Therefore other procedures; solvent gradient chromatography, chromatotron, selective solvent extraction and medium pressure liquid chromatography (MPLC) were employed. MPLC was the most successful technique and flabelliferins could be separated from the most complex mixtures and also from a specific florescent flabelliferin-binding agent. However, depending on flabelliferin profile, other techniques had specific value.The flabelliferin FB (antimicrobial triglycoside) and the flabelliferin FD (inactive diglycoside) and three new flabelliferins (FN, triglycoside of MW884; FE, diglycoside of MW 738 and FF, monoglucoside of MW579) were separated from pooled specimens of PFP collected from Ampara, Anamaduwa, Polonnaruwa and Mannar. Starting from 200g. of PFP, 88.8 mg of FB,52.3 mg of FD, 7.8 mg of FE30.6 mg of FF and 5.6 mg of FN were isolated by an MPLC technique. Starting from 200g of PFP from Jaffna, 300mg of F-II was isolated using a chromatotron. Selective solvent extraction using ethyl acetate from a methanol extract is a simple method of isolating FD from flabelliferin profiles containing triglycosides and tetraglycosides. A technique based on direct MPLC (without mentanol extraction, petroleum ether cleaning, acetone extraction and dry cellulose chromatography was worked out to separate not only the flabelliferins in their pure state but also the carotenoids and the free sugars in PFP. This has the advantage of not subjecting materials to heat and usage of a lesser amount of chemicals. In addition it is less time consuming and gives a yield 2.4 fold that of the indirect method of isolation. The flabelliferins were hydrolyzed by trifluoro acetic acid (0.1M) and the aglycone was confirmed as á-sitosterol by GC/EI/MS studies of the trimethyl silyl derivative and also by 1H-NMR and 13C-NMR.Elucidation of the structure of FB was important due to its potential for use as an antimicrobial agent. The FB was analyzed by Hakomori method followed by GC/EI/MS and also by NMR. It showed that the sugar chain was linked to á- sitosterol by the anomeric C of glucose (áconfiguration), which is linked to two rhamnoses (both were of a anomeric configuration) 1?2 and 1?4 linkages. FB therefore has a branched glycosidic moiety. Similarly, the structure of FD (diglycoside) showed that the sugar chain was linked to á- sitosterol by the anomeric C of glucose. Glucose is linked to a rhamnose by a-1?linkages. The anomeric C of glucose (á configuration) was linked to the á- sistosterol FF (monoglucoside). Through the linkage positions of FE, and FN could not be determined due to insufficient data. It was found that in FN(MW 884), the carbohydrate moiety consists of two glucoses and a rhamnose with a glucose terminus. The sugar moiety is attached to the sapogenin (á- sitosterol) by the other glucose. Similarly, FE(MW 738) showed a carbohydrate moiety of two glucoses. The last part of the study concerned the effect of debittering with naringinase on the nutritive status of ICR mice. Results showed that incorporation of bitter PFP at 10 percent level in WHO recommended standard rat/mice breeding feed, rediced weight gain significantly compared to the control (p=0.029). Debittering of PFP reversed the effect (p=0.88) with respect to control. As debittering not only hydrolyzed F-II but also FB, tjos was mpt [rppf tjat F-II was the causative agent for reducing weight gain. Using data from the morphological study, two special natural PFP's were detected. One contained F-II but not FB(bitter PFP) and the other contained only FB (bitter PFP) and the other contained only FB and not F-II (non-bitter PFP). Results showed that the bitter PFP reduced weight gain compard to control (p=0.021) but the non-bitter increased weight gain compared to control (p=0.014). It is concluded that, provided F-II is absent, PFP at 10 percent level does not have a weight reducing effect. On the contrary it appears to be growth promoting. This may be due to its carotenoid content.Item Studies on the pectic polysaccarides of wattakka cultivar (Cucurbita spp.) and some applications in food formulations(University of Sri Jayawardenepura: USJ(Main)., 2004) HERATH, HMTIn this study, pectin from the fruits of Cucurbita spp, namely, Wattakka, Meemini and Butternut cultivara, was investigated as a potential souce for use in food industry. Towards this end, basic and applied research on the following work was carried out. (a) Isolation and characterization of the cell wall polysaccharides of Cucurbita moschata, Watakka cultivar, using non-degradative methods. The native pectin was purified and characterized to obtain its monimer sugar composition. (b) A comparison was also made on the non-starch polysaccharide compositions of the three cultivars of Cucurbita spp. As components of dietary fibre. (c) Extraction of pectin from the three cultivars of Cucurbita spp. Using an industrial process and determination of chemical and physical characteristics and (d) a study of the applications of pectin from the Wattakka cultivar in some food formulations. The pectic polysaccharides of the cell walls of Wattakka were isolated as alcohol insoluble residue using a modified method described by Ng et al. (1998). This alcohol insoluble residue, was sequentially extracted, under non-degradative methods, using CDTA (0.05 M, pH6.5) at 20§ - 22§ C, for 6 h (CDTA-1) and 2 h (CDTA-2), respectively. The residue thus obtained was further extracted with Na2CO3-2), respectively. The extracts of all fractions were filtered, neutralized and dialysed exhaustively and freeze dried. The monomer sugar compositions of the fractions wre determined after 12 M H2SO4 hydrolysis, neutralization and deivatization to alditol acetates and quantified by GC techniques. The uronic acid content was determined calorimetrically.The yield of the cell wall material of the edible portion of the Wattakka fruit was 29.6 percent, (as alcohol insoluble material), consisting of 91 percent carbohydrates, mainly pectic polysaccharides. The total CDTA and Na2CO3 fractions accounted for 24.7 percent and 3.7 percent respectively, of the original material. The monomer sugar composition of the fractions, showed that the branching points of pectic material (as indicated by the ratio of rhamnose; uronic acid) were high in CDTA-1 (1:38) and CDTA-2 fractions (1:30), while the Na2CO3-1 (1:22) and Na2CO3-2 (1:25) fractions contained less branching points. The major neutral sugar in all fractions was galactose , which was a significant characteristic of Cucurbita spp. The main fraction of the cell wall CDTA-1, was subjected to anion exchange chromatography (DEAE- Trisacryl column), more or less a homogenous individual polymer was obtained, when eluted with 0.25 M NaCI. This polymer contained 84 percent of uronic acid, with a rhamnose to uronic acid ratio of 1:44. The major neutral sugar of the polymer was galactose, with substantial amounts of arabinose and mannose. The non-starch polysaccharides (NSP) of all three cultivars were determined as dietary fibre. Wattakka contained 11.8 g/100g of NSP, of which 22 percent was soluble fibre. The NSP of the other two cultivars was less, Meemini containing 9.1g/100g (20 percent soluble fibre) and Butternut containing 8.9g/ 100g (18 percent soluble fibre). NSP of Cucurbita spp. May be of importance in terms of its physiological role in human systems. The pectin from all three cultivars of Cucurbita spp. Was extracted under industrial conditions (in HC1 at pH 1.3 for 2h, followed by alcohol precipitation), after pre-processing using three different methods. The yields of the pectin were in the range of 7.4 to 28.8 percent for all three cultivars and was found to be dependent on the method of pre-processing. These yields are in the same range of values reported for citrus ( 25 percent) and apple 15-18 percent). The chemical characterization of the industrially estracted Wattakka pectin was classified as a high mehoxyl (HM) pectin, as it had methoxyl content of over 7 percent. However, the Wattakka pectin showed weak gelling properties (gel grade 100) as compared to citrus pectin (gel grade 150), with a low rupture point (0.7K Pa) and a low compression ability (18 N). The presence of higher concentrations of neutral sugars in the pectic polymer may hinder the gel forming ability. Both other cultivars, Meemini and Butternut, did not demonstrate gel forming characteristics, probably due the high acetyl content (over 2 percent). Hence it could be concluded that the use of these two cultivars in food applications are limited. Wattakka pectin had an average molecular weight of 2927 daltons and its viscosity properties increasing markedly at a pH of 4.6 at a concentration of 1 percent (w/v). Wattakka pectin was applied to several food formulations and its technological characteristics were investigated as compared to that of citrus pectin. Wattakka pectin was incorporated into a dietetic fruit juice, showed a higher viscosity values (18-30 cP) as compared to citrus pectin (12-17 cP), at concentrations of 0.15-0.25 percent (w/v), imparting a body and texture to the product. Wattakka pectin when added to a formulation of stirred yoghurt (non-fat), demonstrated the formation of a stabilized milk-pectin complex by showing an increase in the viscosity, ranging from 2200 to 3800 cP, at pH 4-45 and at concentrations ranging from 0.15 - 0.25 percen (w/v). These stabilizing properties, however, were not demonstrated when Wattakka pectin was used in fat contining stirred yoghurt formulations. When Wattakka pectin was used in emulsions of oil-in-pectin solutions, an emulsion stability of 100 percent was obtained at a concentration of 2 percent (w/v), as compared to cirtrus pectin which showed a 40 per cent stability similar conditions.Item Analysis of water quality in well water in the vicinity of an industrial zone and analysis of sources and effects(University of Sri Jayawardenepura: USJ(Main)., 2002) UHANOWITA, SPThe Ratmalana-Moratuwa area located in the south of Colombo has developed as an industrial residential suburb of Colombo in the 1950's. It has been revealed that the environmental problems in the area are very acute due to the activities of large number of industries. About 150 industries are densely packed, within this area. Most of these industries are non-agro based industries they can be categorized in to several groups such as paint industries, galvanizing industries, asbestos industries, dying and finishing , pharmaceutical industries and battery manufacturing industries. According to an investigation of discharges produced in the area it was found that major part of the effluent was released via surface drainage to Lunawa and Bolgoda Lake. The main issue associated with this industrial pollution is water contamination. Even though many studies have been carried out on surface water contamination, studies on ground water contamination have not been done in detail. The intention of this study was to analyze ground water samples at various sites of the industrial zone and to relate the results to the present industrial activities of the area. From these studies it was found that the chemical ozygen demand (COD) levels far exceed the acceptable portable water quality standards, indicating high chemical pollution. In addition it was found that Chemical pollutants are present in the ground water. Biological oxygen demand (BOD) has exceeded the tolerant limits for portable water level at several locations. The trace metals that exceed the levels are Zn, Fe,Cd, Pb and Ni which had not been noted in a previous study done by De Silva etal.(1986). Low pH values in some of the samples indicate water contamination with acids. Some samples have been contaminated with acids and it was revealed by low pH of some samples. Most industries in the area dispose their effluents directly in to the drainage system without any prior treatments. Ground water contamination is thought to be a direct consequence of these inefficient effluent systems. This study discusses the analytical results of water samples taken from various locations of the industrial area and describes the industrial activities in the area that might have caused this ground water contamination, and discusses possible health effects due to use of polluted water as a source of potable water as a source of potable water in this well water.Item Chemical studies and standardisation of Ayurvedic drugs(University of Sri Jayawardenepura: USJ(Main)., 1986) FERNANDO, ISFAn approach developed to standardize commercially available complex Ayurvedic drugs with particular reference to the application of both classical and instrumental analytical techniques is presented.Navaratna and Buddharaja Kalkas were studied and the monographs giving standards in terms of identity, quality and purity were compiled for their 34 raw materials. Standard samples of the two kakas were prepared, the preparative methodologies were discussed and the standards for the two compound preparations were specified. Umbelliferone was identified as a marker compound to establish the presence of asafoetida in Navaratha Kalkaya. Use of GLC profiles to establish the presence or absence of the other ingredients in Navaratna Kalkaya was discussed. Finally the results obtained for the two standard samples were compared with those of commercial products.Item Some microbiological studies of the milk-curd of Sri Lanka(University of Sri Jayawardenepura: USJ(Main]., 1981) ABEYAGOONASEKERA, YHAn investigation carried out to identify the micro-organisms present in milk-curd transported to Colombo from different areas of Sri Lanka revealed the invariable presence of streptococci, lactobacilli and yeast in all the samples tested. All bacterial strains isolated were able to curdle milk in pre cultures with an accompanying change in the pH reached by streptococci and lactstreptococci and lactobacilli in curd also changes with the maturation of the curd indicating the higher acid tolerance of the lactobacilli.Identification experiments done a s described by M. Elisabeth Sharpe, revealed that all bacterial strains belonged to the group of lactic-acid-bacteria. Those identified were two strains of Streptoccoccus lactis, four strains of Thermobacteria and two strains of Betabacterian. When pure cultures of streptococci and lactobacilli were inoculated into milk or into an artificial medium of pH 6.8-7.0, the lactobacilli showed a prolonged initial lag period which was not observed with streptococci. This and similar observations led to the assumption that the lactobacilli were relatively more acidophilic compared to the streptococci. The two bacterial types differed in their preferred pH range and always occurred together in commercially available curds. The efficiency of acid production in milk, by Streptococcus lactis and the thermobacterial strain (B) were tested separately and in combination to investigate whether there was a cumulative effect. However, neither there was a cumulative effect. However, neither a stimulatory nor an inhibitory effect on each others rates of acid production was observed when they were grown in combination. The methods of E.W. Beech et al. (4) were used in the identification of yeasts present in milk. These identifications were confirmed with the help of the Food Research Institute, Colney Lane, Norwich, U.K. Strains identified were Candia parapsilosis and C.Krusei. None of the yeast strains grew in pure cultures in milk, other than when in combination with bacteria. It seems to be that milk alone does not provide a suitable medium for the growth of the yeasts. The yeasts did not ferment lactose and perhaps they depend on bacterial hydrolysis of lactose or other compound s for their carbon source, However, even in the absence of bacteria the yeast strains were capable of growing in milk to some extent if the pH of the milk was lowered. The growth of yeasts together with a Streptococcus lactis strain apparently had very little effect on enhancing the growth of bacteria. However, the growth of yeasts in combination with the thermobacteria apparently had a stimulating effect on the growth of the bacteria. This possibility, however has to be investigated further. The rates of alcohol production of the yeasts isolated were studied using an Eubilliometer. Significant differences in the efficiency of alcohol production by the different yeasts were observed. An analysis of variance of the data confirmed that a significant difference exists at least between some of the types tested. Experiments were carried out to isolate and identify as many as possible different flavour compounds occurring in the milk-curds using curd samples prepared with S.lactis and thermobacteria. Gas-licuid chromatographic analysis of the flavour extracts showed that ethnol and ethylaectate were present in both extracts. Thin layer chromatographic analysis revealed the presence of butanal, propanone and ethanal, and the absence of methanal pentanal and butanone in the flavour extracts. It was also observed that some of the flavour compounds differed quantitatively in extracts made with Streptococcus lactis and thermobacteria. A consumer preference/difference test was carried out with a taste-panel using curds prepared by a Streptococcus lactis strain and a thermobacterial strain. The tast-panel was of unanimous opinion that there was a difference between the curd samples. However, as the normally available curds contain both bacteria and yeasts a Ranking test was also comducted with the taste-panel using curds prepared with different combinations of the isolated micro-organisms. The results indicated that the taste-panel preferred the laboratory-prepared curd, using different combinations of the isolated micro-organisms to the commercially prepared curd, and also that the best curd sample was prepared by using a single bacterial species viz. Streptococcus lactis.Item Content, bioavailability, bioconversion and antioxidant activities of carotenoids in some Sri Lankan fruits and green leafy vegetables(University of Sri Jayawardenepura: USJ(MED)., 2004) CHANDRIKA, UGThe aim of this study was to evaluate the content and bioavailability/bioconversion of pro-vitamin A carotenoids in selected fruits and green leafy vegetables (GLV) from Sri Lanka and to elucidate the effects of different processing and preparation methods. The fruits used in this study was papaya, palmyrah, jak, lavalu and beal. GLV used in this study were kathurumurunga (Sesbania grandiflora). Mukunuwanna (Alternathera sesessilis) gotukola (Centella asoatica). Manioc (Manihot esculenta), sarana (Triathema konogyna), nivithi (Spinacea oleracea) and and thampala (Amaranthus caudatus). In the caase of papaya, palmyrah and jakfruits content of caroenoids were determines using MPLC and visible spectrophotometry and in the case of lavalu and GLV by HPLC method. The vitamin A and á- carotene status in Wistar rats was assessed with and without fat in the diet in the case of papya and mukunuwann leaves using HPLC method. An in vitro method that simulates human digestion was used to measure the fraction of pro-vitamin A carotenoids that is release for absorption (bioaccessible) from selected GLV using HPLC method. Separation and quantification of the carotenoids in two major varieties of Carica papaya grown in Sri Lanka indicated that red- and yellow-fleshed varieties had different carotenoid profiles. Yellow-fleshed papaya contained three major carotenoids. i.e. á-carotene. ?cryptoxanthin and C-carotene. In addition to these three carotenoids, red-fleshed papaya also contained lycopene and á-carotene-5,6 epoxide. It is interesting to note that red-fleshed papaya has significantly higher á-carotene and lycopene than yellow -fleshed variety. Hence the calculated mean retinol equivalent (RE) was 506.7 ñ 54.9 ægkg-1 fresh weight (FW) in yellow -fleshed papaya, whereas in red-fleshed papaya it was 202 ñ 48 ægkg-1 FW. Study with Wistar rats has shown that notwithstanding the colour of the flesh a significant increase of serum vitamin A is shown only if papaya is fed incorporated to standard diet (p\<0.001) rather than given separately (with out incorporated to standard diet ). Red-fleshed variety shows higher (66.7ñ 4.2 æg/g) á-carotene levels in the liver compared to the yellow fleshed variety (5.4ñ 1.0 æg/g. p\<0.001). Six carotenoids were detected in jackfruit kernel. The carotenes are á-carotene. a-carotene. á-zeacarotene. a-zeacarotene, á-carotene 5, 6 epoxide and a dicarboxylic carotenoid crocetin were identified. This corresponds theortically 25.5 RE/100g FW. Our study indicated that jackfruit is a souce of pro-vitamin A carotenoids, but not as good as papaya. Serum retinol concentrations of rats supplemented with jackfruit kernel shown to be significantly higher (p=0.008) compared with control group. The same is true for liver retinol (p=0.006). Quantification was carried out by RP-HPLC. This results shows biological conversion of pro-vitamin A in jackfruit kernel appears satisfactory. Thus increased consumption of ripe hak fruits could be advocated as a part of a strategy to prevent and control of vitamin A deficiency in Sri Lanka. Four major carotenoids were detected in palmyrah fruit pulp (PFP). The carotenoids are a-carotene, ?-carotene. Lycopene and á-zeacarotene and this corresponds to 22.3 RE/100g. FW. The results of in vivo bioavailability and bioconversion studied showed that pro-vitamin A carotenoids from PFP were also bioavailable and bioconvertible. The major carotenoid in lavalu (Chrysophyllum roxhburgii) was trans-violaxanthin (113 mg.kg-1.63 percent). Also present was cisviolaxanthin ( 19 percent) neoxanthin (3 percent). ?-cryptoxanthin monoepoxide ( 11percent) lutein á-cryptoxanthin, ?-carotene and á-carotene. The retinol equivalent of the pulp was only 13.8 RE/100g FW. The study shows that lavalu is not a good source of pro vitamin A. Further the structural properties of the carotenoids make bioconversion studies with animals or humans futile. So that the best use of lavalu carotenoids is as a food colourent for oil based foods.In the case of beli (Aegle marmelos) there were only trace amount of caroenoids and cannot be used as a pro vitamin A supplement. The all-trans á-carotene content in fresh GLV ranged from 63.1 æg/g FW in leaves of mukunuwanna to 133.8 æg/g FW in the thampala. Cooking of fresh leaves resulted in some losses of all-trans-á-carotene and retention ganging from 56 to 76 pecent in the malluma and 27 to 73 percent in the fried preparation. Cooking with water or coconut milk resulted in retentions ranging from 36 to 88 percent . The in vitro accessibility of all-trans-á-carotene in cooked vegetables ranged from 14 to 43 percent in malluma, and from 12 to 36 percent in the fried preparation. In the GLV cooked with coconut milk the in vitro accessibility ranged from 12 to 26 percent compared with 4 to 8 percent when cooked with water. The á-carotene content, retention and in vitro accessibility widely varied between different varieties of green leaves. The three types of different traditional cooking procedures show about the same retention. Preparations with oil scraped coconut and coconut milk had improved in vitro á-carotene accessibility. In conclusion, the green leafy vegetables and fruits studied have variable contents and in vitro accessibility of pro-vitamin A carotenoids and thus contribute in various degrees to recommended daily allowance (RDA) of vitamin A. For green leafy vegetables such as kathurumurunga, mukunuwanna, manioc, gotukola sarana, thamplala an adequate contribution to RDA of vitamin A can be obtained by consumption of green leafy vegetables (100g, FW) with high pro-vitamin A content judging from in vitro accessibility and traditional prepatation methods of Sri Lanka with addition of coconut, coconut milk or coconut oil. None of fruits studied would cover the total vitamin A requirement if consumed in normal amounts. Papaya, jack and palmyrah should be considered as good sources of pro-vitamin A carotenoids, as they would be a sound vitamin A contributing complement to the diet. This study also revealed that some fruits such as lavalu and beli are not good sources of pro-vitamin A as believed by the general population. These findings can be useful in dietary intervention programmes to alleviate vitamin A deficiency in Sri Lanka as well as other developing countries.Item A Study of Tuberculosis in the Galle health division -Sri Lanka(University of Sri Jayawardenepura: USJ(MED)., 1981) MATHILAL, SSNo AbstractItem Study on the chemistry and bio activity of some medicinal plants of Sri Lanka(University of Sri Jayawardenepura: USJ(Main)., 1990) SUMATHIPALA, MPRCentella asiatica (Umbelliferae) and Aegle marmelos (Rutaceae) are widely used as a vegetable and a fruit respectively. These two plants are used in Ayurvedic and indigenous medicinal systems in Sri Lanka.Earlier researchers have reported the effect of aqueous extracts of these two plants on the classical and alternative pathways of activation of human complement in vitro. Activity direvted fractionation of the aqueous extracts indicated that most of the activity was condined to the polysaccharide fraction. This thesis reports the preliminary chemical analysis of the polysaccharide fraction of Centella asiatica and Aegle marmelos. These polysaccharides were hydrolysed and methanolysed. T.L.C and H.P.L.C. analysis were done for analyzing their monomer composition. The immunomodulating activity and biological activity of these fractions using the Artemia bio assay were studied.Item Modeling an epileptic brain using discrete and continuous neural network models(University of Sri Jayawardenepura: USJ(MED)., 2004) SELVARAJAN, SVarious neural network models on epileptic behavior of the human brain were investigated. Several experimental studies have been carried out to simulate behavior of human epileptic brain recently. One of such experiment is the study carried out by Schiff etal. To study the firing behavior of neural networks in hippocampus slices of rat brain. In that study human epileptic brain activity was introduced using the high potassium concentration ([K+]?), where slices from the hippocampus of the temporal lobe of rat brain where exposed to artificial cerebrospinal fluid. Before introducing the high potassium concentration, it was observed that signals recorded from brain slices contained no spikes while with the introduction of high potassium concentration, spikes appeared in random intervals as in the case of brain having epilepsy. The brain-slice experiment mentioned above has been examined using both discrete and continuous neural network models. This discrete model was based on the model developed by Biswal et.al. and Dasgupta et.al. (BD model). In this model, the effect of high potassium medium was introduced through a Hebbian learning mechanism which is switched on during the simulation under reduced inhibition. The sub-passes which play a crucial role in reproducing experimental results in BD model are found to be not necessary when random weights at different stages of the simulation are introduced. A continuous biophysical neural network model was also developed to describe the outcome of the brain-slice experiment mentioned above. In addition, effect of the input current and the potassium concentration changes on dynamics of a single neuron and population of neurons were investigated. It was found that the maps of network activities exhibits stable stationary states and bursting states like trajectories similar to those were found in experiments on hippocampus slices. The discrete and continuous neural network models developed in this wirk were able to successfully reproduced the experimental results.Item The Glycamic indices of different varieties of rice in Sri Lanka(University of Sri Jayawardenepura: USJ(MED)., 2000) HETTIARACHCHI, PGlycaemic index is an important indicator of the physiological effect of consumption of food. It is defined as the glycaemic response after consumption of a 50g carbohydrate portion of food expressed as a percentage on that of an equal carbohydrate portion of a standard, usally white bread or glucose. The glycaemic index indicates how rapidly the bliid glucose levels change on consumption of different foods containing carbohydrates. Hence glycaemic index is beneficial in the management of diabetes mellitus, coronary heart disease, obesity as well as in sports medicine. In Sri Lanka since rice (Oriza sativa) is the staple food, estimating glycaemic indices of different varieties of rice is considered important. If the glycaemic index of a specific variety of rice is not known, advice given to patients with diabetes may not be evidence based. The selection of a variety of rice to be consumed by patients with diabetes mellitus should be based on the physiological effect of the rice consumed. The main objective of this study was to determine the glycaemic indices of different varieties of rice in Sri Lanka. Eight varieties of paddy (red/white pericarp) were obtained. Steam parboiling of four varieties were carried out. Dehusking and milling of samples were carried out. Bread was obtained from the Industrial Technology Institute. The Soxhlet extraction of fat, Kjeldhal method for determination of nitrogen content, Juliano's Method for amylase and enzymatic hydrolysis of dietary fiber of rice flour were carried out. Digestible carbohydrate content of rice and bread were determined by enzymatic hydrolysis and Nelson's method. Drying at 100øc to constant weight in an oven for moisture content of rice and bread and the Dean and Stark method for moisture of cooked rice were carried out. Twenty two healthy adults of both sexes aged between 25-50 years employed in a fibre mill in Udugampola-Gampaha district were selected on a volunteer basis among 38 subjects. Informed consent was obtained from subjects and glucose tolerance test was performed to exclude diabetes. Rice and bread portions containing 50g carbohydrates were fed tio22 human subjects following an overnight fast. Fasting blood samples followed by half hourly samples for two hours were drawn. Blood glucose was determined by the glucose oxidase/ peroxidase method using an autoanalyser. The area under the blood glucose curve (AUC) for varieties of rice for each subject was calculated. Average AUC of 3 values for bread was calculated. Glycaemic indices of different varieties of rice were determined. The glycaemic indices of carieties of rice varied between 55-73, and Bg 350 had the lowest glycaemic index. There was no significant difference between glycaemic indices of varieties of white raw and some varieties of red raw rice. The chemical composition of varieties of rice did not show a significant correlation with the glycaemic indices. Parboiling Bg 94 - 1, BW, Bg 351, increased the soluble content. A low glycaemic index was identified in varieties of red parboiled rice (BW 272 6B, Bg 351) and Bg 350. the glycaemic index could not be predicted from the colour of the rice grain nor the chemical composition. Red parboiled varieties and Bg 350 could be recommended for patients with diabetes.Item Effect of chromium supplementation on glycemic status and serum lipid concentrations in type 2 diabetes mellitus(University of Sri Jayawardenepura: USJ(MED)., 2005) JAYARATNE, SABackground: Chromium is a micronutrient, considered to be essential for birnak carbohydrate and lipid metabolism. Several studies have demonstrated the beneficial effect if chromium on the glycemic status and lipid profile of type 2 diabetes mellitus. Objective: To study the effect of chromium supplementation on glycemic status and serum lipid concentrations in type 2 diabetes mellitus. Method: Sixty type 2 diabetic patients were randomly assigned to two groups. A double blind technique was used and the patients received either 250æg of chromium picolinate or placebo for 12 weeks. The fasting blood glucose, glycated haemoglobin, total cholesterol, high density lipoprotein cholesterol, low density lipoprotein cholesterol and triacylglycerol concentrations were measured at baseline and 2,4,6,8,10 and 12 weeks after initiation of treatment. Serum insulin concentration was also measured at baseline and 12 weeks after initiation of treatment. Results: The fasting blood glucose, glycated haemoglobin, total cholesterol, low density lipoprotein cholesterol and triacylglycerol concentrations did not show a significant difference between the two groups during or at the end of the trial. High density lipoprotein cholesterol is the only parameter that showed a significant increase in concentration starting at 8 weeks after the initiation of treatment, and this trend continued until the end of 12 weeks (p=0.005). Serum insulin resistance calculated from the insulin concentration also did not show a significant difference between the two groups. Conclusion: Results of the trial shows a statistically significant improvement in the high density lipoprotein cholesterol concentration of the supplemented group when compared with the baseline value (p=0.0005). But parallel comparison of the high density lipoprotein cholesterol concentration of the two groups at the 4th, 8th, and 12th week did not show a significant difference. Supplementation of chromium had no effect on the glycemic status and the other lipid parameters.Item Photochemical and pharmacological studies on some medicinal plants of Sri Lanka(University of Sri Jayawardenepura: ., 1985) RATNAYAKE, CKThis thesis covers the phytochemical studies and some biological studies of two medicinal plants belonging to the family Apocynaceae namely, Alstonia macnophylla and Alstonia scholanis. The second part of this thesis describes the estimation of vasicine content in Adhatoda vasica and its seasonal variation in the plant. The second chapter describes the taxonomy and chemical characteristics of the genus Alstonia. The biosynthesis of indole alkaloids including the formation of strictosidine and the plausible biogenetic pathways of some Alstonia alkaloids are given in detail. General methods of extraction of alkaloids that have been used so far, their advantages, disadvantages and more advanced and improved techniques are discussed in the fifth chapter. According to the pharmacological reports on Alstonia alkaloids, it is evident that the two plants we have studied have significant medicinal value. In the first part of the eight chapter, the method of extraction, fractionation and isolation of alkaloids of A. macnophylla and A.scholanis described in detail including the precautions undertaken to isolate alkaloids in their genuine form. Structure of twelve alkaloids have been elucidated from these two plants. One of which is a new alkaloid, 19-hydroxyvincama jine. In these studies we have found that local A.macnophylla is different from the same species found in other countries due to the presence of talcarpine, vincamajine, 19-hydroxyvinvama jine, vincorine and cabucraline. Talcarpine and picraline deacetyl are reported for the first time from the genus Alstonia. These studies give us significant evidences for chemotaxonomic relationship among different species of Alstonis and some other genera such as 7abennemontana, Stnychons, pleiocanpa and Aspidospenma. The effect of alkaloidal extracts of these two plants (leaf and stem bark) on alternative and classical pathways of human complement was tested in vitro. These findings suggest that these two plant alkaloids can be used as immunomodulating agents. Seasonal variation of vasicine content in five parts of A.vasica was studied. According to these studies vasicine content is highest during the month of August. Therefore it is advisable to collect that plant materials between the months of July to September for preparation of Ayurvedic medicine in order to obtain the highest amount of vasicine which is the most active ingredient in the plant A.vasica.Item Development of RT-PCR for rapid detection of dengue virus type 1-4 from clinical specimens and a preliminary phylogenetic study of dengue virus isolates from Sri Lanka(University of Sri Jayawardenepura: USJ(MED)., 2004) VELATHANTHIRI, VGNSDengue virus is the causative agent of dengue fever (DF) and its complications; dengue haemorrhagic fever (DHF) and dengue shock syndrome (DSS). It has four serotypes (Dengue 1-4) and is a member of the family flaviridae. Ot is one of the important causes of morbidity and mortality through out the sybtropical and tropical regions, including Sri Lanka. 1. Patients with dengue need to be closely monitored for evidence of haemorrhage and shock. In order to do this, it is necessary to differentiate patients with dengue from non dengue patients. This however, is a challenge since dengue often presents with non specific symptoms such as fever, headache and body aches. Therefore, it requires an aetiological diagnosis based on laboratory confirmation of disease. In this regard, an ideal diagnostic tool should be sensitive, specific, reliable, rapid cheap technically less demanding and it should also be able to detect dengue in the early stages of the disease. Detection of the circulating serotype is very important in epidemiological surveillance of the disease. Laboratory diagnosis of dengue infection is normally based on detection of dengue virus specific antibodies and isolation of dengue virus from patient's serum. Serological diagnosis has been proven to be of less value in the early stages of illness. Current technologies for isolation and identification of the dengue virus are based on complicated systems such as suckling mouse brain or mosquito inoculation or cell culture technique. These methods are technically demanding, expensive and time consuming. The availability of molecular diagnostics such as the Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) technique has made it possible to perform rapid detection of the viral RNA. In this study, I present an alternative design of the RT-PCR technique that can detect and simultaneously identify the sero type of the dengue virus. I designed sets of primers and developed an RT-PCR for rapid detection and simultaneous identification of dengue virus and its serotypes in cell culture supernatant and in serum samples. It was evaluated for sensitivity, specificity and compared with other standard laboratory diagnostic methods. The test is based on 5 sets of primer pairs specific for dengue viruses within the non-structural (NS) 5 region of the dengue virus genome. A universal primer set that would bind to target sequence shared by all the four serotypes of the virus within the NS5 region, are used. The resulting PCR products are detected by gel electrophoresis and staining with ethidium bromide. The RT-PCR was developed with serotype specific primers, which were also designed within the NS5 region of each serotype for the identification of dengue serotypes. The amplified products of different sizes were obtained with different dengue serotypes and were detected by gel electrophoresis. The RT-PCR technique is simple and rapid, capable of not only detecting the dengue virus but also identifying its serotype in clinical specimens. The RT-PCR protocol developed by me was shown to more sensitive than virus isolation in cell culture and equally sensitive in detecting dengue virus and its serotypes in serum specimens;. It was also shown not to cross react with other flaviviruses. In the preliminary phylogenetic study, I compared the nucleotide sequence homology of the Sri Lankan dengue virus isolates of the present study with dengue viruses isolated form other parts of the world. Nucleotide sequence analysis was performed by an automated nucleic acid sequencer on 14 dengue virus isolates (13 dengue type 2 and one dengue type 3). The dengue 2 viruses were most closely related to dengue 2 virus isolated in Sri Lanka and Seychelle in 1990 and 1976 respectively. The dengue 3 virus was most closely related to dengue 3 viruses recovered in Sri Lanka in 1991, 1985 and 1981. Our results suggested that these dengue virus serotypes are evolving locally and any introduced strains failing to become established.Item An Investigation of palmyrah fruit pulp mediated inhibition of intestinal glucose uptake and its toxicity(University of Sri Jayawardenepura: USJ(MED)., 2005) ULUWADUGE, DIReduced weight gain by Institute of Cancer Research (ICR) mice had been previously reported after feeding 10 percent palmyrah (Borassus flabellifer) fruit pulp (PFP) containing feed. In those studies F-II, a steroidal tetraglycoside of á-sitosterol (MW,1030) was strongly implicated as the active principle. The objective of this study was to confirm the above, its effect on glucose uptake and determine the mechanism of action and some of the factors affecting this effect.The study has provided evidence that confirms the weight reducing property of PFP and also demonstrated its ability to inhibit intestinal glucose uptake in mice. Studies showed that feeding of PFP had no effect on fasting blood sugar levels of mice (p=0.64) but significantly reduced the liver glycogen content of test mice (p\<0.01, 11percent). A mixture of flabelliferins (dose: 10mg/50g mouse) extracted by the methods described previously on administration to mice after a glucose challenge resulted in a decline in serum glucose levels (p\<0.0001, 43 percent ), increased intestinal glucose levels (p\<10-10, 83 percent) and increased faecal glucose levels (p\<0.01, 29 percent). On isolating F-II and other flabelliferins (FB, FD/FE) it was found that only F-II has the ability to lower serum glucose (p\<0.001, 43 percent) and increase intestinal glucose (p\<0.001, 50 percent). This effect was shown at a dose of 1mg F-II/50g mouse. The findings of the mice study were supported by evidence from a study conducted with mild diabetic type-II patients. In the human study it was decided to use pinatty, a dries PFP which can be classified as fruit leather. The dose of 6g of pinattu (30mg FII) was calculated on the basis of IC50 studies and mean human body weight. When pinattu was given to mild, type-II diabetic patients who are not under any therapeutic regimen there was a reduction of 24-48 percent serum glucoafter a glucose challenge (post prandial) when compared with their control (post prandial) values. It has also been shown that the inhibition of intestinal glucose uptake by PFP is mediated mainly by F-II and not by the fibre it contains. A fibre control used in the human study showed that a small part of this is due to fibres (2-11 percent reduction in serum glucose level) present in PFP. It has been known for some time that all flabelliferins are bound by a UV active binder. The present study revealed that this binder was comprised of a major carotenoid component identified as phytofluene and a minor component phytoene. Removal of the binder increased the efficiency with which F-II inhibited intestinal glucose uptake even when the dose was reduced to 2 mg mixed flabelliferin/50g mouse. Inhibition of Na+/K+ ATPase activity was found to be a mechanism by which F-II mediates its inhibitory action on intestinal glucose uptake (the IC50 values were 9x10-5M and 5x10-5 M with and without UV binder respectively). Computational studies showed that the lowering of the inhibition by the binder was probably due to the distortion of the carbohydrate moiety of the F-II, which is strongly connected to the activity the molecule. The beneficial effects of any pharmaceutical agent depend on it being non toxic. Results showed that feeding 10 percent PFP containing feed for one week (short term) or feeding 10percent and 50 percent PFP containing feed for 30 days (long term) did not show any increase in the levels of key hepatic enzyme viz ; alkaline phosphatase, alanine transaminase and aspartate transaminase in the test group when compared with that of the controls. Haematological parameters tested (haemoglobin level, packed cell volume, red cell count and white cell count) were not significantly different in the test and control groups. Creatinine levels in the test and control groups were not significantly different from each other indicating no renal damage. Microscopic examination of liver, kidney and intestine of test group did not show any pathological changes when compared to those in the control group. From the overall results it may be concluded that PFP possibly has an application in treating type-II diabetics most probably those that are obese. However the dosage of PFP pinattu must be closely monitored as different PFPs contain different amount of F-II.Item Low level exposure to organophosphate pesticides in the Udawalawe area and its effects on cardio-respiratory and neurological functions.(University of Sri Jayawardenepura: USJ(MED)., 2003) PEIRIS-JOHN, RJOver 90 percent pesticides used today are insecticides, and approximately half of them are organophosphates (OP). OP compounds are anticholinesterases that inhibit cholinesterase activity resulting in increased availability of acetylcholine (Ach) at its sites of release. ACh influences endocrine and exocrine secretions, cardio-respiratory function, the gut and its secretions, muscle power and functions of the brain. Adverse health effects due to its accumulation are multiple. This study investigated the effects of long term and acute occupational and environmental exposure to Ops on the cardiovascular, respiratory and neurological functions.An analytic, follow-up study was conducted among 150 farmers trained in integrated pest management (IPM farmers), 150 untrained farmers (non-IPM farmers) who were occupationally exposed to OP, all of whom were resident in the Uda Walawe irrigation scheme in Sri Lanka. They were evaluate at three tine points, twice during two cultivation seasons and once during a non-exposure season. Fifty marine fishermen living in Moratuwa, away from agricultural areas were evaluated as a control group. A structured interviewer administered questionnaire was used to obtain information on personal details, occupational history, pesticide handling, past-medical history, social history and symptom profile. A medical examination was conducted at each tike point and included, measurement of blood pressure, handgrip force, and evaluation of vibration sense, two-point discrimination and higher mental function. Haemoglobin corrected red blood cell acetylcholinesterase (AChE) activity was measured and ECG recordings were obtained from all subjects. Electrophysiological, neuromuscular transmission and lung function tests were conducted on a randomly selected subset of 30 farmers, 30 freswater fishermen and on the 50 marine fishermen. Data were entered in EpiInfo version 6 (CDC, Atlanta) and analysed using SPSS version 10 (Chicago, Illinois) statistical package. Ethical clearance was obtained from the Ethical Review Committee of the Faculty of Medical Sciences, University of Sri Jayewardemepura. The farmers and freshwater fishermen had, on average, in their lifetime been directly exposed to OP pesticides for 15.5 and 3.86 years, respectively. IPM farmers wore protective clothing during work in the field significantly more than non-IPM farmers (p\<0.05). Personal protective equipment such as goggles, footwear, gloves or masks were rarely used by both groups of farmers. IPM farmers spent considerably less tine per hectare on spraying insecticides as compared to non-IPM farmers (p\<0.001). The mean (ñSD) baseline AChE levels (U/g) during the non-exposure season in the farmers as a single group, in the non-IPM farmers, IPM farmers, in farmers in the freshwater fishermen and in the controls were 29.25(4.05), 29.16(3.81), 29.22(3.99), 27.75(3.35) and 32.53(3.97), respectively. AChE inhibition was significant in the farmers and the freshwater fishermen during both exposure seasons studied (p=0.001). Although IPM farmers spent less time s[raying, and used more protective clothing as compared to non-IPM farmers (p\<0.001), AChE inhibition was similar in the two groups. AChE inhibition in occupational exposure correlated with adverse effects on sensory and motor systems, but not with nerve conduction studies, cardiovascular or respiratory functions. AChE inhibition in environmental exposure did not correlate with any adverse effect detected. AChE inhibition following a single low level exposure persisted for more than 5 days.Symptoms associated with CNS effects predominated following long term and acute occupational exposure to OP. Acute environmental exposure to OP pesticides resulted in symptoms suggestive of CNS effects, which were less prevalent following term environmental exposure.Handgrip force and vibration sense were significantly weaker in the farmers, following long term and acute occupational exposure as compared to the controls (p\<0.05). Vibration sense was significantly weaker in the freshwater fishermen following acute exposure to OP as compared to the controls (p\<0.05). Attention span, short term memory and subtraction skills were weaker in the farmers (both IPM and non-IPM), and freshwater fishermen following long term and acute OP exposure as compared to the controls (p\<0.05). Sensory conduction velocity (SCV) was significantly faster following long term occupational (p=0.001) and environmental (p\<0.001) exposure, and motor conduction velocity (MCV) was significantly slower (p\<0.05) following long term occupational exposure, as compared to the controls. Both SCV and MCV decreased following acute occupational and environmental exposure to OP. Neuromuscular transmission was significantly weaker in the farmers, and the freshwater fishermen, as compared to the controls, following both, long term and acute low level exposure (p\<0.01). Diastolic blood pressure was significantly lower following long term and acute, occupational and environmental exposure to OP(p\<0.005). Systolic blood pressure was significantly lower following acute occupational exposure (p\<0.001), and long term and acute environmental exposure to OP (p\<0.005). The PR-interval decreased following long term and acute, occupational and environmental exposure to OP pesticides (p\<0.05). The QT-interval was significantly shorter during acute occupational exposure to OP(p\<0.001). The observed FVC (p=0.001), FEV1 (P=0.003) and PEF (p=0.035) in farmers during the non-exposure season were significantly lower than those in the controls. There were no significant differences in FVC and FEVI between the freshwater fishermen during both, the exposure and non-exposure seasons, and the controls. Lung function did not correlate with cardiovascular findings following low level OP exposure.Occupational and environmental exposure to OP pesticides in the Uda Walawe irrigation scheme resulted in adverse effects on the cardiovascular, respiratory and nervous systems. Some effects due to environmental exposure are as severe as those due to occupational exposure. The need for close monitoring of OP use and further studies minimize the adverse impacts of OPs is strongly recommended.