The effect of caffeic acid phenethyl ester (CAPE) on metabolic enzymes including acetylcholinesterase,butyrylcholinesterase, glutathione S-transferase,lactoperoxidase, and carbonic anhydrase isoenzymes I,II, IX,and XII

Caffeic acid phenethyl ester (CAPE) is an active component of honeybee propolis extracts. Carbonic anhydrases (CAs, EC 4.2.1.1) are widespread and intensively studied metalloenzymes present in higher vertebrates including humans as many diverse isoforms. Acetylcholinesterase (AChE) is responsible for acetyl choline (ACh) hydrolysis and plays a fundamental role in nerve impulse transmission by terminating the action of the ACh neurotransmitter at cholinergic synapses and neuromuscular junctions. Butyrylcholinesterase (BChE) is another enzyme abundantly present in the liver and released into blood in a soluble form. Lactoperoxidase (LPO) is an enzyme involved in fighting pathogenic microorganisms whereas glutathione S-transferases (GSTs) are dimeric proteins present both in prokaryotic and eukaryotic organisms and involved in cellular detoxification mechanisms. In the present study, the inhibition effect of CAPE on human carbonic anhydrase (hCA) isoforms I, II, IX, and XII, AChE, BChE, LPO, and GST was evaluated. CAPE inhibited these enzymes with K_is in the range between micromolar to picomolar. The best inhibitory effect was observed against AChE and BChE.     

Isolation,technological characterization and in vitro probiotic evaluation of Lactococcus strains from traditional Turkish skin bag Tulum cheeses

Purpose

The present study was undertaken to evaluate in vitro prerequisite probiotic and technological characteristics of ten Lactococcus strains isolated from traditional goat skin bags of Tulum cheeses from the Central Taurus mountain range in Turkey.

Methods

All isolates were identified based on the nucleotide sequences of the 16S rRNA gene. Eight isolates belonged to Lactococcus lactis and two belonged to Lactococcus garvieae. Probiotic potential was determined from resistance to acid and bile salt, resistance to gastric and pancreatic juices, resistance to antibiotic, auto-aggregation, co-aggregation, diacetyl, hydrogen peroxide and exopolysaccharide productions. Technological properties were verified by alcohol, NaCl and hydrogen peroxide resistance and temperature tests.

Results

L. lactis NTH7 displayed high growth at all alcohol concentrations while L. lactis NTH4 grew very well even at NaCl concentrations of 10%. All strains showed to some extent resistance to acid and bile. Five strains exhibited desirable survival in gastric juice (pH 2.0), while three strains survived in pancreatic juice (pH 8.0). All Lactococcus isolates were sensitive to ampicillin, chloramphenicol, erythromycin, vancomycin, kanamycin, gentamycin and tetracycline. Also, only L. lactis NTH7 from among the isolates showed resistance against penicillin. L. lactis NTH10 and L. lactis NTH7 had higher auto-aggregation values in comparison with all other strains. All the strains demonstrated a co-aggregation ability against model food pathogens, particularly, L. lactis NTH10 which showed a superior ability with L. monocytogenes. All the ten strains produced H₂O₂ and exopolysaccharide (EPS); however, diacetyl production was detected for only four strains including L. lactis NTH10.

Conclusion

These results demonstrate that the L. lactis NTH10 isolate could be regarded as a favorable probiotic candidate for future in vivo studies.

     

The Genotoxic Effect of the New Acaricide Etoxazole

Etoxazole is a member of the diphenyl oxazoline class of insecticide, which was newly developed for use on pome fruits, cotton and strawberries as an acaricide. In the present study, genotoxic effects of acaricide etoxazole (ETX) (miticide/ovicide) were investigated using chromosome aberration (CA) test, sister chromatid exchange (SCE) test, and micronucleus test in human lymphocytes. ETX induced the CAs at all concentrations (5, 10, and 20 g/ml) for 24 h and also induced the CA at the highest concentration (20 g/ml) for 48 h only. The inducing the CAs for 48 h treatment period was dose-dependent. In addition, it induced the SCE at all concentrations and treatment periods in a dose-dependent manner as well. Although ETX decreased the mitotic index (MI) at all concentrations and treatment periods dose-dependently, it did not decrease the replication index (RI) when compared to the negative and solvent controls. In addition, ETX induced the micronucleus at all concentrations except 5 g/ml for 48 h. This inducing was dose-dependent as well. It can be concluded that ETX has a potential genotoxic effects in cultured human peripheral lymphocytes.     

Molecular analysis of 16 Turkish families with DHPR deficiency using denaturing gradient gel electrophoresis (DGGE)

Dihydropteridine reductase (DHPR) catalyses the conversion of quinonoid dihydrobiopterin (qBH2) to tetrahydrobiopterin (BH4), which serves as the obligatory cofactor for the aromatic amino acid hydroxylases. DHPR deficiency, caused by mutations in the QDPR gene, results in hyperphenylalaninemia and deficiency of various neurotransmitters in the central nervous system, with severe neurological symptoms as a consequence. We have studied, at the clinical and molecular levels, 17 patients belonging to 16 Turkish families with DHPR deficiency. The patients were detected at neonatal screening for hyperphenylalaninemia or upon the development of neurological symptoms. To identify the disease causing molecular defects, we developed a sensitive screening method that rapidly scans the entire open reading frame and all splice sites of the QDPR gene. This method combines PCR amplification and "GC-clamping" of each of the seven exonic regions of QDPR, resolution of mutations by denaturing gradient gel electrophoresis (DGGE), and identification of mutations by direct sequence analysis. A total of ten different mutations were identified, of which three are known (G23D, Y150C, R221X) and the remaining are novel (G17R, G18D, W35fs, Q66R, W90X, S97fs and G149R). Six of these mutations are missense variants, two are nonsense mutations, and two are frameshift mutations. All patients had homoallelic genotypes, which allowed the establishment of genotype-phenotype associations. Our findings suggest that DGGE is a fast and efficient method for detection of mutations in the QDPR gene, which may be useful for confirmatory DNA-based diagnosis, genetic counselling and prenatal diagnosis in DHPR deficiency.     

A preliminary study of human paraoxonase and PON 1 L/M55–PON 1 Q/R 192 polymorphisms in Turkish patients with coronary artery disease

Paraoxonase 1 (PON 1) is a high‐density lipoprotein (HDL)‐associated enzyme with antioxidant function protecting low‐density lipoprotein (LDL) from oxidation. PON 1 has two amino acid polymorphisms in coding region; L/M 55 and Q/R 192. These polymorphisms modulate paraoxonase activity of the enzyme. PON 1 activity decreases in coronary artery disease (CAD). In the present study, distribution of PON 1 L/M 55 and Q/R 192 polymorphisms and the effect of these polymorphisms on the activities of PON 1, and on the severity of CAD in 277 CAD (+) patient and 92 CAD (?) subjects were examined. PON 1 L/M 55 and Q/R 192 genotypes were determined by PCR, RFLP and agarose gel electrophoresis techniques. Genotype distributions and allele frequencies for PON 1 Q/R 192 polymorphism were not significantly different between controls and CAD (+) patient group (p > 0.05), but in genotype and allele distribution of PON 1 L/M55 polymorphism, there was significantly difference among groups (p < 0.05). Genotype distributions for both polymorphisms were not significantly different between subgroups of single‐vessel disease (SVD), double‐vessel disease (DVD) and triple‐vessel disease (TVD). Serum PON 1 activity was lower in CAD (+) group than in controls and this was also statistically significant (p < 0.001). In both groups, the highest PON activities were detected in LL and RR genotypes. In summary, our results suggest that there is an association between the PON 1 L/M 55 polymorphism of paraoxonase and CAD in Turkish patients but not with PON 1 Q/R 192 polymorphism. However, it is hard to correlate these polymorphisms and severity of CAD. Copyright © 2009 John Wiley & Sons, Ltd.     

Comparison of two different cryopreservation protocols for freezing goat semen

In this study, two different semen cryopreservation protocols were compared to freeze goat semen. The ejaculates (n = 12) were collected by using electro-ejaculator from six mature bucks (two ejaculates per each buck). Each ejaculate was divided into two groups as Protocol 1 (P1) and Protocol 2 (P2). In P1, semen was diluted directly in an extender containing 15% egg yolk, 300 mM Tris, 28 mM glucose, 95 mM citric acid 5% glycerol to a concentration of 200 × 10⁶ sperm/mL. In P2, after the removal of seminal plasma by centrifugation, the semen sample was diluted with the first portion of milk extender consist of 100 mg/mL skimmed milk powder and 27.75 mM glucose (without glycerol) to a concentration of 400 × 10⁶ sperm/mL. The second portion of the milk extender containing 14% glycerol was added to semen gradually in order to achieve sperm concentration 200 × 10⁶ sperm/mL and 7% glycerol level in the final volume. Extended semen was loaded in 0.25 mL straws, held for 2 h at 4 °C, frozen in nitrogen vapor and stored in liquid nitrogen. Post-thaw motility and live sperm rate (mean ± SEM) were significantly lower (P < 0.05) in P1 as compared to P2 (47.50 ± 1.23% vs. 55.63 ± 1.72%; 80.04 ± 1.29% vs. 84.04 ± 1.08%, respectively). However, live intact, total intact, abnormal, reacted acrosome and DNA damaged sperm rates were similar (P > 0.05) in both protocols. It was concluded that both protocols used in this study provided reasonable post-thaw parameters; however, P2 yielded better motility and live sperm rate compared to P1.     

Age,growth and sexual development of solenette,Buglossidium luteum (Risso, 1810), in the central Aegean Sea

Age, growth, spawning period and maturity of the solenette (Buglossidium luteum Risso, 1810) were studied in the central Aegean Sea to provide fisheries managers with essential data for science‐based management. A total of 1220 samples were collected by trawl hauls from July 2004 to June 2007 in ?zmir Bay (Turkey). Sample sizes ranging from 5.3 to 11.6 cm total length were composed of 46% females, 32% males and 22% immature individuals, with a female to male ratio of 1 : 0.7. Age composition stages of the females were from I to IV, and males between I and III. The length–weight relationship was calculated as W = 0.0101L^3.008 for all samples. Estimated von Bertalanffy growth parameters were L = 13.30 cm, t_o = ?0.440 year and k = 0.481 year^?1, with a growth performance index of 1.93 (?’). The spawning period began in April and continued until July. Lengths at first maturity of females and males were 8.1 and 7.9 cm total length, respectively. Both sexes matured at the age of 2 years.