Emin Sofic

Amperometric determination of bonded glucose with an MnO2 and glucose oxidase bulk-modified screen-printed electrode using flow-injection analysis

A screen-printed amperometric biosensor based on carbon ink double bulk-modified with MnO(2) as a mediator and glucose oxidase as a biocomponent was investigated for its ability to serve as a detector for bonded glucose in different compounds, such as cellobiose, saccharose, (-)-4-nitrophenyl-beta-d-glucopyranoside, as well as in beer samples by flow-injection analysis (FIA). The biosensor could be operated under physiological conditions (0.1M phosphate buffer, pH 7.5) and exhibited good reproducibility and stability. Bonded glucose was released with glucosidase in solution, and the free glucose was detected with the modified screen-printed electrode (SPE). The release of glucose by the aid of glucosidase from cellobiose, saccharose and (-)-4-nitrophenyl-beta-d-glucopyranoside in solution showed that stoichiometric quantities of free glucose could be monitored in all three cases. The linear range of the amperometric response of the biosensor in the FIA-mode flow rate 0.2mLmin(-1), injection volume 0.25mL, operation potential 0.48V versus Ag/AgCl) extends from 11 to 13,900mumolL(-1) glucose in free form. The limit of detection (3sigma) is 1mumolL(-1) glucose. A concentration of 100mumolL(-1) yields a relative standard deviation of approximately 7% with five injections. These values correspond to the same concentrations of bonded glucose supposed that it is liberated quantitatively (incubation for 2h with glucosidase). Bonded glucose could be determined in beer samples using the same assay. The results corresponded very well with the reference procedure.

Reduced Brain Antioxidant Capacity in Rat Models of Betacytotoxic-Induced Experimental Sporadic Alzheimer’s Disease and Diabetes Mellitus

It is believed that oxidative stress (OS) plays a central role in the pathogenesis of metabolic diseases like diabetes mellitus (DM) and its complications (like peripheral neuropathy) as well as in neurodegenerative disorders like sporadic Alzheimer’s disease (sAD). Representative experimental models of these diseases are streptozotocin (STZ)-induced diabetic rats and STZ-intracerebroventricularly (STZ-icv) treated rats, in which antioxidant capacity (AC) against peroxyl (ORAC-ROO•) and hydroxyl (ORAC-OH•) free radicals (FR) was measured in three different brain regions: the hippocampus (HPC), the cerebellum (CB), and the brain stem (BS) by means of oxygen radical absorbance capacity (ORAC) assay. In the brain of both STZ-induced diabetic and STZ-icv treated rats decreased AC has been found demonstrating regionally specific distribution. In the diabetic rats these abnormalities were not associated with the development of peripheral diabetic neuropathy (PDN). Also, these abnormalities were not prevented by the intracerebroventricularly (icv) pretreatment of glucose transport inhibitor 5–thio-d-glucose (TG) in the STZ-icv treated rats, suggesting different mechanism of STZ-induced central effects from those at the periphery. Similarities of the OS alterations in the brain of STZ-icv rats and humans with sAD could be useful in the search for the new drugs in the treatment of sAD that have antioxidant activity. In the STZ-induced diabetic animals the existence of PDN was tested by the paw pressure test, 3xa0weeks following the diabetes induction. Mechanical nociceptive thresholds were measured three times at 10–min intervals by applying increased pressure to the hind paw until the paw-withdrawal or overt struggling was elicited. Only those diabetic animals which demonstrated decreased withdrawal threshold values in comparison with the control non-diabetic animals (C) were considered to have developed the PDN.

Antioxidant capacity in postmortem brain tissues of Parkinson’s and Alzheimer’s diseases

Oxidative stress has been associated with damage and progressive cell death that occurs in neurodegenerative disorders such as Parkinsons disease (PD) and Alzheimers disease (AD). The aim of this study was to investigate the antioxidant capacity in postmortem motor cortex (MC), nucleus caudatus (NC), gyrus temporalis (GT) and substantia nigra (SN) from controls (C) and patients with PD and AD. The initial samples consisted of 68 subjects of PD, AD and C. Brains were matched for age, sex and postmortem time. Brain tissue was homogenized in a phosphate buffer pH 7.3 and separated with two-step centrifugation at 15,000rpm for 30 min and 15,000 rpm for 10 min at 4 degrees C. Antioxidant capacity in the supernatants was measured using the oxygen radical absorbance assay (ORAC). The results showed that in the SN of parkinsonians brain the balance between production of free radicals and the neutralization by a complex antioxidant system is disturbed. No changes in the antioxidant capacity of postmortem MC and NC of parkinsonians brain in comparison with C were found. In the SN of parkinsonians brain, antioxidant capacity seems to be lower in comparison with C (p < 0.05). Antioxidant capacity against peroxyl radical showed that MC of AD patients was lower than in the MC of C (p < 0.005). In NC of AD patients the antioxidant capacity against hydroxyl radical was increased in comparison with C (p < 0.04). No changes in the antioxidant capacity were found in brain tissues of AD in comparison with C, when CuSO4 was used as a free radical generator.

Serum nitric oxide concentrations in patients with multiple sclerosis and patients with epilepsy

Nitric oxide (NO), a neurotransmitter and a free radical, has been purported to be involved in numerous neurological diseases. We investigated the serum nitric oxide concentration in 30 patients with multiple sclerosis (MS), in 30 patients with epilepsy and in 30 control subjects. The aim was also to determine whether a statistically significant difference in serum NO concentrations exists between the groups of interest. The total serum nitric oxide concentration was measured using the Griess reaction after reducing nitrates to nitrites with elemental zinc. In the group multiple sclerosis, the mean NO concentrations were Xxa0±xa0SEMxa0=xa031.02xa0±xa01.79xa0μmol/l, in the control group Xxa0±xa0SEMxa0=xa025.31xa0±xa01.44xa0μmol/l and in the group epilepsy Xxa0±xa0SEMxa0=xa022.51xa0±xa01.28xa0μmol/l. Student’s t test showed a statistically significant difference between subjects with multiple sclerosis and the control group (pxa0=xa00.013), as well as between the groups multiple sclerosis and epilepsy (pxa0=xa00.0002). This data confirms that NO may play an important role in the pathogenesis of multiple sclerosis, whereas its role in epilepsy still remains unclear.

Brain antioxidant capacity in rat models of betacytotoxic-induced experimental sporadic Alzheimer’s disease and diabetes mellitus

It is believed that oxidative stress plays a central role in the pathogenesis of metabolic diseases like diabetes mellitus (DM) and its complications (like peripheral neuropathy) as well as in neurodegenerative disorders like sporadic Alzheimers disease (sAD). Representative experimental models of these diseases are streptozotocin (STZ)-induced diabetic rats and STZ-intracerebroventricularly (STZ-icv) treated rats, in which antioxidant capacity against peroxyl (ORAC(-ROO)*) and hydroxyl (ORAC(-OH)*) free radical was measured in three different brain regions (hippocampus, cerebellum, and brain stem) by means of oxygen radical absorbance capacity (ORAC) assay. In the brain of both STZ-induced diabetic and STZ-icv treated rats decreased antioxidant capacity has been found demonstrating regionally specific distribution. In the diabetic rats these abnormalities were not associated with the development of peripheral diabetic neuropathy. Also, these abnormalities were not prevented by the icv pretreatment of glucose transport inhibitor 5-thio-D-glucose in the STZ-icv treated rats, suggesting different mechanism for STZ-induced central effects from those at the periphery. Similarities in the oxidative stress alterations in the brain of STZ-icv rats and humans with sAD could be useful in the search for new drugs in the treatment of sAD that have antioxidant activity.

Brain catalase in the streptozotocin-rat model of sporadic Alzheimer’s disease treated with the iron chelator–monoamine oxidase inhibitor, M30

Low intracerebroventricular (icv) doses of streptozotocin (STZ) produce regionally specific brain neurochemical changes in rats that are similar to those found in the brain of patients with sporadic Alzheimer’s disease (sAD). Since oxidative stress is thought to be one of the major pathologic processes in sAD, catalase (CAT) activity was estimated in the regional brain tissue of animals treated intracerebroventricularly with STZ and the multitarget iron chelator, antioxidant and MAO-inhibitor M30 [5-(N-methyl-N-propargylaminomethyl)-8-hydroxyquinoline]. Five-day oral pre-treatment of adult male Wistar rats with 10xa0mg/kg/dayxa0M30 dose was followed by a single injection of STZ (1xa0mg/kg, icv). CAT activity was measured colorimetrically in the hippocampus (HPC), brain stem (BS) and cerebellum (CB) of the control, STZ-, M30- and STZxa0+xa0M30-treated rats, respectively, 4xa0weeks after the STZ treatment. STZ-treated rats demonstrated significantly lower CAT activity in all three brain regions in comparison to the controls (pxa0<xa00.05 for BS and CB, pxa0<xa00.01 for HPC). M30 pre-treatment of the control rats did not influence the CAT activity in HPC and CB, but significantly increased it in BS (pxa0<xa00.05). M30 pre-treatment of STZ-treated rats significantly increased CAT activity in the HPC in comparison to the STZ treatment alone (pxa0<xa00.05) and normalized to the control values. These findings are in line with the assumption that reactive oxygen species contribute to the pathogenesis of STZ in a rat model of sAD and indicate that multifunctional iron chelators such as M30 might also have beneficial effects in this non-transgenic sAD model.

Elevated serum homocysteine level is not associated with serum C-reactive protein in patients with probable Alzheimer’s disease

AbstractnElevated plasma homocysteine (Hcy) levels have been associated with Alzheimer’s disease (AD) and cognitive impairment. Studies have shown that Hcy may have direct and indirect neurotoxicity effects. The aim of the study was to investigate serum Hcy concentration in patients with probable AD with age-matched controls and to determine whether there was an association between serum Hcy and C-reactive protein concentration in patients with probable AD. We also aimed to determine whether there was an association between serum tHcy concentration and cognitive impairment in patients with probable AD. Serum concentration of total Hcy was determined by the fluorescence polarization immunoassay on the AxSYM system, and serum C-reactive protein (CRP) concentration was determined by means of particle-enhanced immunonephelometry with the use of BN II analyzer. Cognitive impairment was tested by the MMSE score. Body mass index (BMI) was calculated for each subject included in the study. Age, systolic and diastolic blood pressure and BMI did not differ significantly between the two groups. Mean serum tHcy concentration in the control group of subjects was 12.60xa0μmol/L, while in patients with probable AD the mean serum tHcy concentration was significantly higher than 16.15xa0μmol/L (pxa0<xa00.01). A significant negative association between serum tHcy concentration and cognitive impairment tested by the MMSE score in patients with probable AD was determined (rxa0=xa0−0.61634; pxa0<xa00.001). Positive, although not significant correlation between CRP and serum tHcy concentrations in patients with AD, was observed. Increased tHcy concentration in patients with probable AD, and the established negative correlation between serum tHcy concentration and cognitive damage tested by MMSE score in the same group of patients, suggests the possible independent role of Hcy in the pathogenesis of AD and cognitive impairment associated with this disease.

Antioxidant capacity in rat brain after ICV treatment with streptozotocin and alloxan — a preliminary study

Intracerebroventricular (icv) administration of betacytotoxic drug streptozotocin (STZ) produces long-term and progressive cognitive deficits in rats, as well as deficits in cerebral glucose and energy metabolism. These changes resemble those found in the brain of patients with sporadic Alzheimer’s disease (sAD), and therefore, STZ-icv treated rats have been proposed as an experimental model of sAD. In this study the antioxidant capacity (AC), using manual oxygen radical absorbance capacity (ORAC) assay, was measured in the rat brain frontoparietal cortex (FC) and brainstem-cerebellum region (BS-CB) after administration of STZ and another betacytotoxic drug alloxan (AL). Region-specific differences of AC were found, which were more expressed when hydroxyl radical (ORAC-OH°) generator was used in the assay. AC against ORAC-OH° was significantly lower in BS-CB than in FC of the control rats. Furthermore, ORAC-OH° significantly decreased in BS-CB 3-months following the icv administration of AL, but significantly increased following the TG+AL combined treatment in comparison with the controls. However, 3-months following the icv treatment of AL combination with a different glucose transport inhbitor, 3-O-methyl-D-glucose, ORAC-OH° values in BS-CB and ORAC-ROO° values in FC were significantly decreased in comparison to the controls. Our results suggest that betacytotoxic-icv treatment alters antioxidant defense systems in the brain, which particularly regarding the STZ-icv treatment, could be a useful tool in search for possible new antioxidant treatments of the neurodegenerative disorders such as sAD.

The Relation of Erythropoietin Towards Hemoglobin and Hematocrit in Varying Degrees of Renal Insufficiency.

Introduction: Hypoxia is a basic stimulant in production of erythropoietin (EPO). The primary function of erythrocytes is the transport of oxygen to tissues. Erythropoietin stimulates erythropoiesis which leads to increased production of erythrocytes- their total mass. This increases the capacity of the blood to carry oxygen, reduces the hypoxic stimulus and provides a negative feedback of stopping EPO production. The aim of this study was to establish a quantitative relationship between the concentration of erythropoietin, hemoglobin and hematocrit in different values of renal insufficiency. Material and methods: The survey was conducted on 562 subjects divided into two groups: with and without renal insufficiency. EPO, hemoglobin, hematocrit, serum creatinine and additional parameters iron, vitamin B12, and folic acid were determined by using immunochemical and spectrophotometric methods and glomerular filtration rate (GFR) was calculated as well. Results: EPO values (median) grow to the first degree of renal insufficiency, as compared to EPO values of healthy subjects, this increase is statistically significant, p=0.002. With further deterioration of renal function the values of EPO between all pathological groups are decreasing, and this decrease is statistically significant between first and second degree of renal insufficiency (RI) p<0.001. In the group of healthy subjects EPO is correlated rho = -0.532, p <0.0005 with hematocrit. The correlations are negative and strong and can be predicted by regression line (EP0 = 41.375- Hct * .649; EPO = 61.41–Hb * 0.355). In the group of subjects with the first degree of renal insufficiency EPO is in correlation with hematocrit rho=-0.574, p<0, 0005. It is also correlated with hemoglobin rho=-0.580, p< 0.0005. The correlation is negative (EP0= 42.168- Hct * 0.678). In the group of subjects with the third degree of renal insufficiency EPO is in correlation with hemoglobin rho=0.257, p=0.028. The correlation is medium strong and positive. In the group of subjects with third and fourth degree of renal insufficiency EPO is not in correlation with hemoglobin and hematocrit p>0.05. Conclusion: Renal dysfunction, depending on the level of RI effects differently on the biosynthesis of EPO in a diseased kidney, and consequently it also has a different effect on biosynthesis of HB in bone marrow and its content in the blood.

Chemical composition of various Ephedra species

The medicinal significance of Ephedra is based on the sympathomimetic properties of ephedrine (E) alkaloids. Pharmacological effects depend on the phytocomposition of individual Ephedra species. The aim of this study was to measure the total alkaloids content (TAC), total phenolics content (TPC), and total flavonoids content (TFC) and determine their relationship in dry herb of Ephedra major, Ephedra distachya subsp. helvetica, Ephedra monosperma, Ephedra fragilis, Ephedra foeminea, Ephedra alata, Ephedra altissima and Ephedra foliata. Nowadays, medicinal use of Ephedrae herba is limited, but the abuse of its psychostimulants is rising. In this study, TAC, TPC and TFC were determined using spectrophotometric methods. For the first time, ultra-performance liquid chromatography with ultraviolet detection (UPLC-UV) was used for separation and quantification of E-type alkaloids of various Ephedra species. The highest TPC and TFC were found in E. alata (53.3 ± 0.1 mg Gallic acid equivalents/g dry weight, 2.8 mg quercetin equivalents/g dry weight, respectively). The total content of E and pseudoephedrine determined by UPLC-UV varied between 20.8 mg/g dry weight (E. distachya subsp. helvetica) and 34.7 mg/g dry weight (E. monosperma). The variable content and ratio between secondary metabolites determined in different Ephedra species reflects their metabolic activities. Utilization of UPLC-UV unveiled that this technique is sensitive, selective, and useful for separation and quantification of different alkaloids in complex biological matrixes. The limit of detection was 5 ng. Application of UPLC-UV can be recommended in quick analyses of E-type alkaloids in forensic medicine and quality control of pharmaceutical preparations.