A processing route for development of hydroxyapatite (Ca10(PO4)6(OH)2 or HAp)-titania (TiO2) hybrid coatings on titanium alloy (Ti6Al4V) has been established. HAp powders of different size and morphology were synthesized by aqueous precipitation techniques using different precursor couples and XRD, SEM and FTIR were performed for complete characterization. Hybrid coatings were then prepared via sol-gel by incorporating presynthesized HAp powders into a titanium-alkoxide dip coating solution. Titania network is formed by hydrolysis and condensation of Ti-isopropoxide (Ti[OCH(CH3)2]4) based sols. The effect of titania sol formulation, specifically the effect of organic solvents on the microstructure of the dip coated films calcined at 500 &ordm / C has been investigated. The coatings exhibit higher tendency for cracking when a high vapor pressure solvent, such as ethanol (C2H5OH) is used causing development of higher macroscopic stresses during evaporation of the sol. Titania sol formulations replacing the solvent with n-propanol (CH3(CH2)2OH) and acetly-acetone (C5H8O) combinations enhanced the microstructural integrity of the coating during evaporation and calcination treatments. Sol-gel processing parameters such as multilayer coating application and withdrawal rate can be employed to change the titania thickness in the range of 0.120 - 1.1 microns and to control the microstructure of HAp-titania hybrid coatings. Slower withdraw rates and multi-layer dip coating lead to coatings more vulnerable to cracking. A high calcination temperature in the range of 400 &ordm / C-600 &ordm / C lead to more cracking due to combined effect of densification originated stresses and thermal stresses upon cooling.

Flavonoids, present in fruits, vegetables and beverages derived from plants, have been described as health-promoting, disease-preventing dietary supplements, and have activity as cancer preventive agents. The cancer protective effects of flavonoids have been attributed to a wide variety of mechanisms, including modulating enzyme activities resulting in the decreased carcinogenicity of xenobiotics. Cytochrome P4501A1 (CYP1A1) is a Phase I enzyme which is known to be involved in the activation of procarcinogens and Glutathione S-Transferase (GST) is a Phase II enzyme which is largely responsible for the detoxification of carcinogens. In this study, it was aimed to investigate the mechanisms of inhibition of CYP1A1 and GST activities of fish by phenolic compounds/flavonoids. Leaping mullet (Liza saliens), captured from highly polluted sites of izmir Bay, expressing high levels of CYP1A, were used in order to investigate these effects. It was demonstrated that all of the phenolic compounds/flavonoids used, exert an inhibitory effect on both CYP1A1 associated 7-Ethoxyresorufin-O-deethylase (EROD) activity and GST activities of fish, although the degree of inhibition was varied with the flavonoid used. Of the flavonoids tested, the most potent inhibitor of CYP1A1 associated EROD activity was found to be quercetin. The potency of the phenolic compounds/flavonoids to inhibit CYP1A1 associated EROD activity follow the sequence of quercetin &gt / resveratrol &gt / naringenin &gt / hesperidin &gt / rutin with IC50 values of 1.32 &micro / M, 3.59 &micro / M, 9.78 &micro / M, 98.5 &micro / M and 0.64 mM respectively. Quercetin, resveratrol, hesperidin and rutin were found to inhibit EROD activity in a competitive manner, on the other hand, naringenin was found to inhibit EROD activity in a non-competitive manner. Inhibition constant (Ki) values of quercetin, resveratrol, naringenin, hesperidin and rutin were calculated from Dixon plots as 0.12 &micro / M, 0.67 &micro / M, 2.63 &micro / M, 18 &micro / M and 0.1 mM, respectively. In the case of GST enzyme, it was demonstrated that all of the phenolic compounds/flavonoids used, exert an inhibitory effect on both total GST and GST-Mu activities of fish. Of the flavonoids tested, the most effective inhibitor of total GST activity was found to be resveratrol. The potency of the phenolic compounds/flavonoids to inhibit total GST activity follow the sequence of resveratrol &gt / quercetin &gt / rutin &gt / naringenin &gt / hesperidin with IC50 values of 7.1 &micro / M, 24.5 &micro / M, 89 &micro / M, 116 &micro / M and 118 &micro / M respectively. Resveratrol, quercetin and hesperidin were found to inhibit total GST activity in a competitive manner, on the other hand, rutin and naringenin were found to inhibit GST activity in a mixed type manner. Ki values of resveratrol, quercetin, hesperidin, naringenin and rutin were calculated from Dixon plots as 3.2 &micro / M, 12.5 &micro / M, 45 &micro / M, 128 &micro / M and 150 &micro / M respectively. In the case of GST-Mu activity, the most potent inhibitor was found to be rutin. The potency of the phenolic compounds/flavonoids to inhibit GST-Mu activity follow the sequence of rutin &gt / resveratrol &gt / quercetin &gt / naringenin &gt / hesperidin with IC50 values of 66.5 &micro / M, 72.3 &micro / M, 113.5 &micro / M, 135.5 &micro / M and 196 &micro / M, respectively. In conclusion, this study indicated that flavonoids were the strong inhibitors of CYP1A1 associated EROD activity and GST activities of mullet liver. The modulation of drug-metabolizing enzymes by flavonoids is important in terms of human health, since these enzymes can activate or inactivate carcinogens. The potential role of xenobiotic metabolizers CYP1 family in the activation of carcinogens and inactivation of chemotherapeutics suggests a potential therapeutic benefits in inhibiting these enzymes. The results of the present study support the hypothesis that flavonoids may be involved in the prevention of malignant transformation, by reducing the formation of carcinogens through inhibition of enzymes such as CYP1A1 which is known to be involved in carcinogen activation.

Energy transfer is a natural phenomena that bases of the photosynthesis. In photosynthesis, chlorophyls and auxlary absorbing dyes units absorb solar energy and transfer the energy to the reaction center where other chemical reactions occur. Studies showed that energy transfer depends strongly on distance and rigidy of the structure. Rigid structures that close in space transfer energy nearly %100 efficiency. Supramolecular chemistry tries to mimic nature and miniurize natural phenomence on to molecular scale. Thus, articifial photosynthesis has been very popular in supramolecular chemistry. A lot of studies have been dedicated to diffent parts of photosynthesis. BODIPY dyes have very well defined photophysical properties that can be used in multichromic systems and designing molecular switching potential tool on in supramolecular chemistry. In this study extended BODIPY dyes constructed on a rigid xanthane scaffold have been deviced for energy transfer.

Turkish red pine is coastal tree and is a drought resistant pine that withstands more aridity and poor soils than most other timber species growing in the same climatic conditions. In Turkey, this species grows in southern and western Anatolia and is also found in the Marmara region. Drought results in a water deficit in plant tissues, which, in turn, can lead to an imbalance in the redox poise of plant cells, and thus inducing oxidative stress in plants. Resistance to conditions associated with oxidative-stress must, in part, rely on endogenous antioxidative defense mechanisms required to maintain cellular homeostasis. Glutathione is one of the major endogenous antioxidants in plants known to play an important role in plant defense mechanisms. Glutathione S-transferase (GST, EC 2.5.1.18) is a GSH dependent detoxifying enzyme in plants, which catalyzes the conjugation of GSH. In this study, we investigated the changes in cytosolic glutathione S-transferase enzyme activity using CDNB as substrate and total thiol amount in Pinus brutia Ten., related to the drought stress during four months, June to September. The osmotic pressure in the needles was also determined as an indirect measure of drought condition. Together with the increase in the temperature values from June to July, GST enzyme activity increased from 15,78 &plusmn / 1,36 &micro / moles min-1 mg protein-1 to 22,91 &plusmn / 1,99 &micro / moles min-1 mg protein-1 which was statistically significant. However in August, GST activity had fallen to 16,54 &plusmn / 1,61 &micro / moles/min/mg protein, which may be because of a local rainfall at the beginning of the August in the sampling area. In September, GST activity significantly increased with respect to June, in accordance with high temperatures. The total thiol amount was not changed significantly during the sampling period. Although there were statistically significant changes in osmotic pressure in the needdles collected during the same sampling period, it did not exactly correlated to the changes in GST activity.

ABSTRACT PURIFICATION AND CHARACTERIZATION OF HEXOKINASE ISOENZYMES FROM Rhizopus oryzae Dedeoglu, Didem MS., Department of Biotechnology Supervisor: Prof.Dr. Haluk Hamamci Co-supervisor: Dr. Seyda A&ccedil / ar February 2007, 116 pages Glycolysis is the central metabolic pathway for living organisms. Its regulation is important for the yield of the end products which are industrially important. These end products, like lactic acid produced by Rhizopus oryzae, are industrially important. Rhizopus oryzae is a filamentous fungus producing lactic acid and ethanol. The lactic acid yield of R. oryzae is low (&amp / #61566 / 70 %) compared to that of lactic acid bacteria (&amp / #61502 / 95 %) still it is noteworthy because R. oryzae produces only the L (+) form of lactic acid which can be metabolized in the human body. The yield of an industrial process should be high for the feasibility of the production of a particular product. If a way can be found increase the flux through the glycolysis the yield of lactic acid may increase as well. Keeping this in mind we wanted to focus on the first step of glycolysis, hexokinase of R. oryzae. Hexokinase catalyzes the reaction that converts glucose to glucose-6-phosphate. In this study for the first time the two isoenzymes of hexokinase of R. oryzae were purified and characterized by biochemically and kinetically Hexokinase has two isoenzymes. The purified enzymes (isoenzymes1 &amp / isoenzymes2) obeyed Michealis-Menten Kinetics. The Km value of purified isoenzyme 1 is 0.16 mM and isoenzyme 2, 0.21 mM at pH 7.70 for glucose. The Km value of isoenzyme1 for fructose was 28.8 mM. Essentially isoenzyme 2 can not utilize fructose. None of the isoenzymes were inhibited by trehalose-6-phophate.The monomer moleculer weight of isoenzymes were estimated SDS PAGE analysis. There were two different values for molecular weight of isoenzmye 1 / 62.9 and 42.5 kDa and two values for isoenzyme 2 / 56.2 and 41.6 kDa

This study was designed to investigate the cytotoxic and antioxidative properties of Aesculus hippocastanum L. (A. hippocastanum) bark extracts. Dried and powdered barks were extracted in ethanol, methanol, water and ethylacetate at a ratio of 1:6 (w/v). Antioxidative capacity of each extract (ethanol, methanol, water and ethylacetate) were determined by their ability to scavenge 1, 1 -diphenyl-2-picryl-hydrazyl radical (DPPH). Effective concentration (EC50) values were calculated as 0.010 mg/mL 0.011 mg/mL, 0.009 and 0.019 mg/mL, respectively for ethanol, methanol, ethylacetate and aqueous extracts. The highest DPPH radical scavenging activity was demonstrated by ethyl acetate among the four bark extracts of A. hippocastanum. Nevertheless, methanol extract was preferred for the separation, identification and further quantification of its phenolic compounds using HPLC method. Analytical and semi&ndash / preparative HPLC methods were applied to qualify and quantify the isolates. Human Myeloid Leukemia (HL - 60) cell line was used as a model system for the proliferation studies. HL - 60 cells were cultured in the presence of various concentrations (0 to 100 &micro / g/mL) of methanol bark extract and, also, with the various concentrations of standard esculetin. HL-60 cell viability was examined by tryphan blue and the metabolism of tetrazolium salt XTT (2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino) -carbonyl]-2H-tetrazolium hydroxide). XTT effective dose (ED50) values for the proliferation studies of methanol extract and standard esculetin were calculated as 56.18 &micro / g/mL and 21.23 &micro / g/mL, respectively. These results suggested that A. hippocastanum methanol bark extract and esculetin could be considered as a potent antioxidant and cytotoxic agent.

Stroke is an important public health problem and the third leading cause of death after coronary heart diesase and all cancers in all over the world. Free radicals and oxidative stress play important role in the pathogenesis of several diseases including atherosclerosis, stroke, cancer, neurodegenerative diseases such as Alzheimer&#039 / s dementia. The activity of paraoxonase (PON1) aganist phenylacetate is known as arylesterase (ARE). Paraoxonase is an esterase associated with high-density lipoprotein (HDL) and contributes to the protective role of this lipoprotein on low-density lipoprotein (LDL) oxidation. Oxidized LDL is known to play a central role in early events in the progression of atherosclerosis which is a risk factor for stroke. Glutathione S-transferases (GSTs) catalyze the conjugation of nonpolar compounds to reduced glutathione (GSH) and detoxify toxic metabolites produced within the cell by oxidative stress to protect cells from oxidant injury. v The maximum ARE enzyme activity was detected at 10 mM Tris-HCl buffer, pH 8.0 and at 45 &deg / C. ARE enzyme was saturated with its substrate phenylacetate around 20 mM concentration. The apparent Km and Vmax values of human blood serum ARE for phenylacetate were found as 1.66 mM and 3300 nmol/min/mg, respectively. The maximum GST enzyme activity was detected at 2 mM potassium phosphate buffer, pH 5.5 and at 65 &deg / C. GST enzyme was saturated with its substrate, CDNB around 4.5 mM concentration and with its cofactor, GSH around 8 mM concentration. The apparent Km and Vmax values of human blood serum GST for CDNB substrate were found as 2.8 mM and 0.43 nmol/min/mg and for GSH were found as 4.11 mM and 0.23 nmol/min/mg, respectively. In addition, effects of three different heavy metal ions, Cd+2, Hg+2 and Ni+2, on human blood serum ARE and GST activity were studied and half maximal inhibitory concentrations (IC50) were determined. The main objective of this study was to investigate the human blood serum ARE and GST activities in patient and control groups using the optimized conditions. For this purpose, blood samples were collected from 172 ischemic stroke patients and 105 controls. Then serum obtained from blood samples were used to determine ARE and GST activities. The mean of ARE activity in patient group (n=172, 109.9 &plusmn / 32.5 U/mL ) was insignificantly lower than the mean of ARE activity in control group (n=105, 113.5 &plusmn / 33.1 U/mL, P=0.284). GST activity of the patients (10.8 &plusmn / 4.4 U/L) was insignificantly higher than that of controls (10.5 &plusmn / 4.2 U/L, P=0.483 ). In addition, statistical analysis showed hypertension, diabetes and HDL as significant risk factors of stroke.

Clinical S. aureus strains were gathered from four hospitals, two in Turkey (Hacettepe hospital 200 strains and Ankara Hospital 106 strains) and the other two from Libya (Aljalla Hospital 88 strains and Jamahyria Hospital 62 strains). The clinical specimens were collected form different sources including blood, urine, wound, pus, burn, sputum, semen, catheter and aspiration. Patients were aged between 0 to 84 years and from both sexes. Resistance to Methicillin was determined by measuring the Oxacillin MIC / this was done by using the oxacillin E-test, with resistance defined as an MIC of &gt / 2 &micro / g ml. In this study all isolates displayed an Oxacillin MIC of &amp / #8805 / 256&micro / g/ml. The MRSA strains were (56%) in Turkish hospitals, and (59%) in Libyan hospitals. The percentage of the VRSA and VISA in Libyan hospitals was (7%) and (26%) respectively, although the percentage of VRSA in Turkish hospitals was only 2% and there were no intermediately susceptible Staphylococcus aureus (VISA). Besides the MRSA isolates, Coagulase Negative Staphylococcus showing Methicillin resistance was collected from clinical isolates in thirteen patients in Turkish hospitals. In both countries, the majority MRSA isolates were multiresistant to more than five classes of antibiotics including / Ampicillin, Amoxicillin, Tetracycline, Erythromycin and Ciprofloxacin. Most of the MRSA isolates were from blood (68%), wounds (57%) and pus (50%).The results of genetic investigations indicated that the mecA gene was present in the majority of isolates in both countries / the community acquired MRSA type (ccr-BIV) was present in three samples out of thirty in Turkish hospitals and in one case out of twenty in Libyan hospitals / There was no case out of fifty specimens that carry the hospitals acquired MRSA type (ccr-BI, II, III) in both countries. Besides the Methicillin resistance gene, the incidence of Tetracycline resistance gene was quite high (tetM and tetK 50%) in Turkish hospitals isolates, and the prevalence of Panton-Valentine Leukocidin gene was high (PVL 70%) in Libyan hospitals specimens.

The main object of the presented study was investigation of the use of cross-linked enzyme (tyrosinase) aggregates (CLTA) obtained from crude mushroom extract for a rapid phenolic content analysis in wines. In addition, a comparison of phenolic characteristics of Turkish red wines was performed. Reproducible and reliable results in total phenolic measurement were obtained with CLTAs similar to pure tyrosinase and tyrosinase obtained from crude mushroom extract. Measurement of total phenolic content is possible both in standard solutions and in complex matrices, such as wine. In a very short time period, 10 seconds, phenolics content in red and white wines produced from grapes of Turkey were investigated by using CLTAs. Results were consistent when compared to a well known phenolic measurement method, Folin-Ciocalteau. CLTAs exhibited very high operational stability and retained more than 90% of its activity after 30th use. Moreover, it showed good shelf-life stability for about 2 months storage by maintaining 90% of its maximum activity. So, use of CLTAs prepared from crude mushroom extract is an effective, fast and cheap alternative in total phenolics measurements in wines. Moreover, a novel catalase phenoloxidase (CATPO) produced by a fungal microorganism, Scytalidium thermophilum, was studied to check its capabilities in phenolics measurements. This novel catalase phenol oxidase showed similarly good results, exhibiting widesubstrate selectivity.

In the polyol pathway, Aldose reductase (AR) is an important enzyme in reduction of aldehydes and aldosugars to their suitable alcohols. AR, using NADPH as a coenzyme, has a molecular weight of 37 000 dalton. AR in its activated form, known to increase the sorbitol accumulation in lens, is responsible for the cataract formation in diabetis diseases. Therefore, the inhibition of aldose reductase is important to prevent the incedence of cataract formation in diabetus mellitus. In the treatment of diabetis dependent cataract, chemically synthetized drugs were sometimes less than beneficial due to the severe side effects they cause. Recently a huge amount of study has been intensified on developing new drugs from natural compounds and even by utilizing plant extracts for their easily metabolizing polyphenolic compounds. In this study, BLAR, source of enzyme, was obtained as crude via differential centrifugation and ammonium sulfate precipitation. The enzyme assay conditions were optimized for the protein, substrate, coenzyme, and salt concentrations, also for the effects of pH and temperature. Ocimum basilicum, Lavandula stoechas, Melissa officinalis, Glycyrrhiza glabra L. and Tilia tomentosa were selected as commonly used alternative medicine plants. Plant extracts were prepared in ethanol and ethyl acetate and their inhibitory effects were tested on crude bovin lens aldose reductase enzyme. Fifty percent inhibitory concentrations (IC50) were found between values of 25.53 &micro / g/mL and 54.15 &micro / g/mL for ethanol extracts and between 41.55 &micro / g/mL and 82.96 &micro / g/mL for the ethyl acetate extracts of selected plants. In addition, the plant extracts were also characterized for their antioxidant activities by of 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging method and test of total phenolic content (TPC) .