Validation of an In-House-Developed GC-MS Method for 5α-Cholestanol According to ISO 15189:2012 Requirements.

OBJECTIVE: The aim of this study was to validate a gas chromatography-mass spectrometry (GC-MS) method for the measurement of 5α-cholestanol in the clinical laboratory in agreement with ISO 15189:2012. MATERIALS AND METHODS: The GC-MS performance was evaluated and proficiency testing data were used to estimate the measurement uncertainty of the method considering the recommendations of international guidelines. RESULTS: The calibration curves were linear from 6 to 50 µmol/L, with r2 >.99. The limit of detection and limit of quantitation were determined to be 0.36 and 2.58 µmol/L, respectively. The bias ranged from -18.9% to 15.2% for 6.5, 18.3, and 66 µmol/L. The intra- and interassay reproducibility was <20% at the various concentrations studied. The expanded uncertainty was determined to be 50.9%. CONCLUSION: The GC-MS method for the measurement of 5α-cholestanol has proved to have acceptable analytical performance for use in the clinical laboratory.

The effects of pre-pregnancy obesity and gestational weight gain on maternal lipid profiles, fatty acids and insulin resistance.

OBJECTIVES: Pregnancy is associated with physiological alterations in insulin sensitivity and lipid metabolism. This study investigates the associations between pregestational body mass index (pBMI) and the rate of gestational weight gain (rGWG) in the second trimester with the biomarkers of lipid, fatty acids metabolism and insulin resistance. METHODS: Sixty nine pregnant women followed. The body weights of the pregnant women were measured and blood samples were obtained at 11-14th and 24-28th weeks of pregnancy. Glucose, total cholesterol, triglyceride, HDL cholesterol, LDL cholesterol, insulin levels and fatty acids were measured. Rate of GWG (kg/week) and The Homeostasis Model Assessment for Insulin Resistance (HOMA-IR) were calculated. The pregnant women were stratified according to their pBMI and the 2nd trimester rGWG. RESULTS: The rate of GWG was significantly higher for the group with pBMI<25, compared to the group with pBMI≥25 (p=0.024). Triglyceride, total cholesterol, LDL and HDL cholesterol were significantly increased in the second trimester compared with the first trimester. Palmitic acid, oleic acid, linoleic acid, myristic acid, docosahexaenoic acid (DHA), arachidonic acid (AA), total omega-6 (n - 6) and omega-3 (n - 3) fatty acid levels and n - 6/n - 3 ratio were significantly higher in the second trimester. Glucose was significantly decreased and insulin was increased in the second trimester. In the overweight/obese group; HOMA-IR, insulin, AA, palmitoleic acid and stearic acid were found to be high in comparison to the group with low/normal pBMI. No parameters were associated with rGWG. CONCLUSIONS: The changes in lipid parameters, free fatty acids, insulin and HOMA-IR in the second trimester were compatible with the changes in lipid metabolism and the development of insulin resistance. Pregestational BMI was shown to have a stronger influence on lipid profile, insulin resistance, and fatty acids than rGWG.

Effect of Blood Cell Subtypes Lysis on Routine Biochemical Tests.

BACKGROUND: The aim of this study is to establish the contribution of blood cells subtypes on hemolysis. METHODS: Separated blood cell subtype suspensions prepared with blood from 10 volunteers were serially diluted to obtain different concentrations of cell suspensions. The cells were fully lysed and cell hemolysates were added (1:20) to aliquots of serum pool. Thus, seven serum pools with different concentrations of interferent were obtained for each blood cell subtype. Biochemical parameters and serum indices were measured by an autoanalyzer. As cell lysis markers, free hemoglobin was measured by spectrophotometry while myeloperoxidase and ᵝ-thromboglobulin were measured by enzyme immunoassay. The percent changes in analyte levels of the serum pools were evaulated by Wilcoxon Signed Rank Test and compared with clinical thresholds defined for each test. RESULTS: The clinical thresholds were exceeded in lactate dehydrogenase, potassium, aspartate aminotransferase, creatine kinase, magnesium, total protein, total cholesterol, inorganic phosphate, glucose for red blood cells (RBC); lactate dehydrogenase, aspartate aminotransferase, total protein, inorganic phosphate and glucose for platelets (PLT). Free hemoglobin was significantly correlated with RBC (r=0.999; p=0.001), while myeloperoxidase and b thromboglobulin showed no significant correlation to white blood cells (WBC) and PLT, respectively. CONCLUSIONS: The effect of RBC hemolysis in serum on the routine biochemical tests are clearly established, yet, additional studies are required in order to verify this kind of effects of PLT and WBC hemolysis.

Investigation of iron's neurotoxicity during cerebral maturation in the neonatal rat model of haemolysis.

INTRODUCTION: Haemolytic disease of newborns due to rhesus and AB0 incompatibility is encountered frequently in neonatal clinics and may lead to severe haemolysis. In this study, it is suggested that important amounts of iron released with haemolysis may have a toxic effect on the brain parenchymal tissue, and the severity of the toxic effect can be correlated with the maturation of the brain barrier systems. To demonstrate the accumulation and the neuro-toxic effects of free iron (Fe) in the brain an experimental haemolysis model with various maturation phases was performed. MATERIAL AND METHODS: The study was composed of 48 Wistar rats with the following ages: five days old (Group A), 10 days old (Group B), and 19 days old (Group C). Each group was divided into three experimental subgroups and three control groups. Experimental groups were treated with intraperitoneal 75 mg/kg/day phenyl hydrazine hydrochloride for haemolysis. RESULTS: We demonstrated that the blood brain barrier (BBB) is permeable in five-day-old newborn rats and is mature in 10- and 19-day-old rats. Iron staining and neuronal damage were detected in group A and group B rats. No damage was detected in the brain tissue of group C animals. The presence of iron staining and neuronal damage in group B with mature BBB may suggest the existence of other incomplete barrier systems different from BBB that lead to iron accumulation in the brain. CONCLUSIONS: Blood brain barrier has a partial role in Fe transport, and the alternative barrier systems may also be involved. It could be supposed that after maturation of all barrier systems, excessive Fe penetration to the brain cannot occur. Our findings showed that the toxic amounts of iron may penetrate into the brain parenchyma of newborns despite the BBB preservation and cause neuronal damage in newborns, but the mature brain is not affected by the same magnitude blood levels.

A multicenter nationwide reference intervals study for common biochemical analytes in Turkey using Abbott analyzers.

BACKGROUND: A nationwide multicenter study was organized to establish reference intervals (RIs) in the Turkish population for 25 commonly tested biochemical analytes and to explore sources of variation in reference values, including regionality. METHODS: Blood samples were collected nationwide in 28 laboratories from the seven regions (≥400 samples/region, 3066 in all). The sera were collectively analyzed in Uludag University in Bursa using Abbott reagents and analyzer. Reference materials were used for standardization of test results. After secondary exclusion using the latent abnormal values exclusion method, RIs were derived by a parametric method employing the modified Box-Cox formula and compared with the RIs by the non-parametric method. Three-level nested ANOVA was used to evaluate variations among sexes, ages and regions. Associations between test results and age, body mass index (BMI) and region were determined by multiple regression analysis (MRA). RESULTS: By ANOVA, differences of reference values among seven regions were significant in none of the 25 analytes. Significant sex-related and age-related differences were observed for 10 and seven analytes, respectively. MRA revealed BMI-related changes in results for uric acid, glucose, triglycerides, high-density lipoprotein (HDL)-cholesterol, alanine aminotransferase, and γ-glutamyltransferase. Their RIs were thus derived by applying stricter criteria excluding individuals with BMI >28 kg/m2. Ranges of RIs by non-parametric method were wider than those by parametric method especially for those analytes affected by BMI. CONCLUSIONS: With the lack of regional differences and the well-standardized status of test results, the RIs derived from this nationwide study can be used for the entire Turkish population.

The effect of malnutrition on protein glycosylation in children.

OBJECTIVE: The goal of this study was to evaluate the effect of protein energy malnutrition on protein glycosylation by investigating transferrin isoform pattern and its relationship to the degree of malnutrition and the biochemical markers of nutritional status in children. METHODS: Forty one children with mild (n=23) and severely/moderately (n=18) acute malnutrition and 29 controls were enrolled in the study. Serum transferrin isoforms were determined by isoelectric focusing electrophoresis. Transferrin, prealbumin, zinc, iron and insulin-like growth factor-1 (IGF-1) were measured using automated analyzers. Findings : Asialotransferrin and disialotransferrin were significantly higher in severely/moderately malnourished patients compared to controls (P=0.04 and P=0.04, respectively). Other transferrin isoform patterns were not different among three groups. Serum IGF-1, transferrin and iron levels of severely/ moderately malnourished group were significantly lower than tose of controls (P=0.001, 0.02 and 0.03, respectively). Serum prealbumin and zinc levels were similar in all three groups. Serum IGF-1, transferrin and iron levels, and all transferrin isoform patterns were not significantly different in mildly malnutrition group from other two groups. CONCLUSION: The changes in transferrin isoform pattern observed in malnourished patients may indicate that malnutrition is a catabolic state which has effects on glycosylation.

The effects of different syringe volume, needle size and sample volume on blood gas analysis in syringes washed with heparin.

INTRODUCTION: We evaluated the effect of different syringe volume, needle size and sample volume on blood gas analysis in syringes washed with heparin. MATERIALS AND METHODS: In this multi-step experimental study, percent dilution ratios (PDRs) and final heparin concentrations (FHCs) were calculated by gravimetric method for determining the effect of syringe volume (1, 2, 5 and 10 mL), needle size (20, 21, 22, 25 and 26 G) and sample volume (0.5, 1, 2, 5 and 10 mL). The effect of different PDRs and FHCs on blood gas and electrolyte parameters were determined. The erroneous results from nonstandardized sampling were evaluated according to RiliBAK's TEa. RESULTS: The increase of PDRs and FHCs was associated with the decrease of syringe volume, the increase of needle size and the decrease of sample volume: from 2.0% and 100 IU/mL in 10 mL-syringe to 7.0% and 351 IU/mL in 1 mL-syringe; from 4.9% and 245 IU/mL in 26G to 7.6% and 380 IU/mL in 20 G with combined 1 mL syringe; from 2.0% and 100 IU/mL in full-filled sample to 34% and 1675 IU/mL in 0.5 mL suctioned sample into 10 mL-syringe. There was no statistical difference in pH; but the percent decreasing in pCO2,, K+, iCa2+, iMg2+; the percent increasing in pO2 and Na+ were statistical significance compared to samples full-filled in syringes. The all changes in pH and pO2 were acceptable; but the changes in pCO2, Na+, K+ and iCa2+ were unacceptable according to TEa limits except fullfilled-syringes. CONCLUSIONS: The changes in PDRs and FHCs due nonstandardized sampling in syringe washed with liquid heparin give rise to erroneous test results for pCO2 and electrolytes.

Health profiles of methyl bromide applicators in greenhouses in Turkey

<p><b>INTRODUCTION</b>Methyl bromide is a toxic substance that has hazardous effects on human health with acute and chronic exposure. Our previous study showed that methyl bromide applicators frequently use large amounts of methyl bromide haphazardly in greenhouses in the prefectures of Narlidere and Balcova in the Aegean city of Izmir. This study aims to evaluate the health conditions of these workers.</p><p><b>MATERIALS AND METHODS</b>Our previous study showed that there are 38 methyl bromide applicators in our study area. After the informed consent of methyl bromide applicators was obtained, a questionnaire was used for a survey of demography and symptoms. Each subject was examined before and after application of the compound. Blood and urine samples were collected and stored. Blood samples were analysed for methyl bromide and bromide ion, kidney and liver function tests and lipid profile.</p><p><b>RESULTS</b>The age range of subjects was 19 to 53 years (mean age: 41 +/- 8.57). This study showed that methyl bromide applicators use large amounts of methyl bromide disregarding legal regulations and that some of them had nonspecific complaints. Subjects had been working as methyl bromide applicators for approximately 9.7 +/- 4.15 years. A total of 69.7% of methyl bromide applicators reported that they did not use protective equipment while 33.3% of them had a history of acute methyl bromide intoxication. A statistically significant relationship was found between the usage of protective equipment and the level of blood bromide ion in the blood (P <0.05).</p><p><b>CONCLUSION</b>Usage of methyl bromide, training, screening and follow-up of applicators must be rigorously controlled in accordance with national legal arrangements and international protocols. Greater efforts are required in the implementation of controls to achieve the targets set by the legal regulations and to ensure continual improvement in the limitation of the risks of this environmental hazard.</p>

Does cisplatin chemotherapy decrease the MDP uptake of normal bone? An experimental study.

OBJECTIVE: Bone scan is the accepted initial imaging modality for skeletal metastases. Cisplatin is a cell-cycle nonspecific antineoplastic agent used in some chemotherapy regimens. Knowing that platinum reacts with phosphate compounds such as methylenediphosphonic acid (MDP), decreases bone resorption and new bone formation, it can be proposed that cisplatin chemotherapy may decrease Tc-99m MDP bone uptake. We aimed to demonstrate, if present, the decrease in bone uptake and to determine the duration of this effect. METHODS: Thirty male Wistar rats were randomized into five groups, namely, placebo group (G1) and cisplatin groups (G2, G3, G4, G5). Pre-therapy bone scintigraphies were obtained in all the groups. Cisplatin chemotherapy was given as infusion. Post-therapy bone scintigraphies were obtained 10 min, 1 h, 24 h, and 72 h after chemotherapy in groups G2-G5, respectively. A placebo bone scintigraphy was obtained 10 min after infusion of serum physiologic in G1. Plasma samples for cisplatin plasma values were obtained. The graphite furnace atomic absorption spectrophotometry technique was used for cisplatin analysis. Quantitative analysis (bone uptake ratios) was performed by drawing regions of interest on the right femur, vertebral column, and adjacent soft tissues. The injection/examination time delay and the net injected MDP doses were also noted. RESULTS: There was no statistically significant difference in bone uptake values, injected MDP doses or injection/examination time delay in any group. Cisplatin plasma values were significantly different in G2, G3, G4, and G5 (P < 0.05) but not in G1. CONCLUSIONS: Cisplatin chemotherapy seems to have no effect on the Tc-99m MDP uptake of normal bone.

Effects of selenium supplementation on antioxidant defense and glucose homeostasis in experimental diabetes mellitus.

The objective of this study was to investigate the effects of different forms of Se supplementation on the antioxidant defense and glucose homeostasis in experimental diabetes. Sodium selenate (SS) or selenomethionine (SM) were administered (2 micromol Se kg(-1) day(-1)) via orogastric route to streptozotocine (STZ)-induced diabetic rats in addition to basal diet for 12 weeks. Glucose levels in whole blood, glutathione peroxidase (GSH-Px) activity in erythrocytes, Se and fructosamine levels in plasma were evaluated monthly. Plasma Se levels increased significantly in all diabetic groups compared to basal measurements, being more prominent in SM group [p(SM(3)/SM(0)) = 0.018]. The increase in GSH-Px activities was significant at the end of the second month in SS [p(SS(2)/SS(0)) = 0.028], whereas at the end of the third month in SM the value was lower [p(SM3/SM0) = 0.018] and the unsupplemented diabetic control (DC) groups, p(DC(3)/DC(0)) = 0.012. Glucose increased significantly only in DC group. Fructosamine increased gradually in all diabetic groups, being significant in DC and SS groups. At the end of the third month, highest fructosamine levels were observed in SS group, which were significantly higher than the SM group [p(SM/SS) = 0.010]. In conclusion, Se augmented the antioxidant defense by increasing GSH-Px activity and this effect was more prominent when Se was supplemented as SM, which exerted positive effects also on glucose homeostasis.

Amniotic fluid ferritin as a marker of intestinal damage in gastroschisis: a time course experimental study.

BACKGROUND/PURPOSE: Intestinal damage (ID) is closely related to morbidity and mortality in gastroschisis. This study was performed to determine the intraamniotic substances that may correlate ID and also to verify their time course levels that would be useful for determining when ID starts in gastroschisis. METHODS: In this study, 13-day-old fertilized chick eggs were used. The amnioallantoic membrane was perforated to create amnioallantoic cavity in all embryos. Gastroschisis was created in gastroschisis group to simulate human gastroschisis. Amnioallantoic fluid samples were collected from the embryos on the 13th to 19th gestational days, and the intestines of each group were harvested for evaluation. Amnioallantoic levels of interleukin-8, ferritin, alkaline phosphatase, and amylase were measured. Serosal thickness of the intestines in each group was evaluated. RESULTS: Increasing amnioallantoic fluid levels of interleukin-8, alkaline phosphatase, and amylase were found in both groups. In contrast to control group, ferritin levels, as a sign of inflammation, were found increased only in gastroschisis group. Histopathologic examination of intestines in the gastroschisis group showed a significant increase in the serosal thickness especially after the 16th day. CONCLUSION: Increases in amnioallantoic fluid levels of ferritin show promise as a marker for determining ID encountered in gastroschisis but warrant further investigation.

In vitro release of elements from prosthodontic base metal alloys: effect of protein-containing biologic environments.

PURPOSE: This study aimed to investigate the release of nickel, copper, zinc, cadmium, magnesium, and lead from prosthodontic base metal alloys into protein-containing biologic solutions. MATERIALS AND METHODS: Dissolution experiments were conducted in either 3% Dulbecco's Modified Eagle Medium or 3% bovine serum albumin solutions for 7 weeks. RESULTS: The protein-containing dissolution solutions and dissolution time did not have a significant effect on release of elements from the alloys (Kruskal-Wallis, P > .05). CONCLUSIONS: The amount of the released elements was well below the dietary intake levels of these elements. This study is important given the widespread use of the base metal alloys and the continuing public concerns/questions regarding the health benefits/risks associated with these materials.

The relationship between trace elements and cardiac markers in acute coronary syndromes.

Previous studies have demonstrated increased serum copper and iron levels and decreased selenium and zinc levels in patients with myocardial infarction. Furthermore, the prognostic value of the levels of trace elements in myocardial infarction has been stressed. We examined serum levels of Cu, Fe, Zn and Se, as well as glutathione peroxidase (GPx), a selenoenzyme with antioxidant properties, and C-reactive protein (CRP), a marker of inflammation, in acute coronary syndromes (ACS) regarding their relationship to cardiac troponins and creatine kinase-MB mass (CK-MBm), important prognostic markers. Serum trace elements, GPx activity and CRP were determined in 70 patients with ACS who were admitted within 12 h after the onset. Differences in these parameters were evaluated in three groups of patients divided according to the levels of cardiac markers: group III consisted of patients with high increases in cTnT, cTnI and CK-MBm (> or =0.9 ng/mL, > or =1.0 ng/mL, > or =30 ng/mL, respectively), patients with milder increases in these markers were included in groups II and I consisted of patients with values just above the upper reference limits. Serum Fe levels increased significantly in group II and even more prominently in group III compared to group I (p = 0.04, 0.002, respectively). There was no significant difference between groups II and III. The increase in serum Cu was significant in group III compared to both groups II and I (p = 0.04, 0.001, respectively). There was no significant difference between groups I and II regarding Cu and Zn. The decrease in serum Se and GPx levels was significant only between groups III and I (p = 0.004 for Se and p = 0.0001 for GPx). CRP levels showed a significant increase in group III compared to groups II and I (p = 0.03 and 0.001). CRP showed a significant positive and GPx a significant negative correlation to the cardiac markers cTnT, cTnI and CK-MBm. Cu was positively correlated to all cardiac markers, while the positive correlation between Fe and cardiac markers was significant only for cTnI. Both Zn and Se were negatively correlated to cTnT, and Se was also to cTnI. In conclusion, the increase in serum levels of Cu and Fe and the decrease in serum levels of Zn and Se in patients with higher levels of troponins and CK-MBm imply that trace element levels are related to the degree of myocardial damage and thus may play a role in the pathogenesis of ischemic heart disease. The strong correlations between cardiac markers and both CRP and GPx suggest that these parameters are promising prognostic factors in acute coronary syndromes.

Effects of selected antibiotics on pancreatitis induced liver and pulmonary injury.

OBJECTIVE: To investigate the protective effect of antibiotherapy in the early phase of acute pancreatitis on cellular injury induced in lungs and liver. BACKGROUND: Cellular viability and plasma nitric oxide (NO) levels were assessed to determine the efficacy of highly bactericidal imipenem and quinolones on liver and lung injury. METHODS: Imipenem, levofloxacin or saline were administered to rats with caerulein induced pancreatitis. Twenty-four hours later serum amylase, aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, and NO levels, pancreatic interstitial inflammation, acinar cell necrosis, acinar cell vacuolisation, peripancreatic fat necrosis; spotty necrosis, focal inflammation of liver and inflammatory processes in the lungs were assessed. RESULTS: Enzyme levels in the antibiotherapy groups were lower than in the control group. Serum NO levels were higher only in the imipenem group. Levofloxacin decreased acinar cell vacuolisation in the pancreas; interstitial edema, neutrophilic infiltration and interstitial enlargement in the lungs. Antibiotherapy decreased spotty necrosis, apoptosis and focal inflammation in the liver. CONCLUSIONS: Although treatment with imipenem is associated with higher NO levels than levofloxacin, levofloxacin prevents organ injury more effectively than imipenem in acute pancreatitis. Our results indicate that antibiotherapy in the early period of necrotizing pancreatitis prevents cellular damage induced in pancreas, liver and lungs.

Hair zinc level in Down syndrome.

Immunological, endocrinological, and haematological abnormalities are relatively common in people with Down syndrome (Cuadrado & Barrena, 1996; Decoq & Vincker, 1995; Hestnes et al., 1991; Sustrova & Strbak, 1994; Nespoli, Burgio, Ugazio & Maccario, 1993; Kempski, Chessells & Reeves, 1997; Kivivuori, Rajantie, & Siimes, 1996; David et al., 1996; Gjertson, Sturm & Berger, 1999). Zinc is one of the elements that act in the maintenance of normal function of these systems. This study was designed to investigate zinc levels in children with Down syndrome. Zinc levels were measured in hair using atomic absorption spectrophotometry. The hair zinc level of 19 children with Down syndrome was compared with the zinc level of 11 typically developing children. Hair zinc levels were found to be significantly lower (p < .05) in those with Down syndrome (average 95.18 +/- 56.10 ppm) than in the typically developing children (average 208.88 +/- 152.37 ppm). Some of the problems experienced by children with Down syndrome may be due to these low zinc levels, but further research is required to confirm these results, and to establish any correlation with these problems.

Cadmium exposure in tobacco workers: possible renal effects.

Cadmium is a nephrotoxic metal widely used in industry and the main source of Cd in general population is smoking. Considering that the source of Cd in cigarettes is the tobacco leaf, the exposure to Cd was evaluated in workers employed at a tobacco leaf processing factory. Blood and urinary Cd levels were measured by flameless atomic absorption spectrometry in 87 workers and 35 controls. Urinary enzymes, total protein, albumin and uric acid were also determined to investigate the possible nephrotoxic effects of Cd. Blood Cd levels were significantly higher in workers (1.63 +/- 1.95 microg/L) than in controls (0.91 +/- 1.15 microg/L) (p = 0.044). The increase observed in urinary Cd levels of workers was non significant (0.56 +/- 0.5 microg/g creatinine in workers and 0.46 +/- 0.5 microg/g creatinine in controls). Both in workers and in controls, subjects smoking >10 cigarettes/day showed significantly increased blood Cd levels compared to non-smokers (p = 0.000 and p = 0.011, respectively). In workers, urinary alkaline phosphatase (ALP), gamma glutamyl transferase (GGT), total protein, and uric acid were observed to be significantly increased (p = 0.013, p = 0.000, p = 0.000, p = 0.025, respectively), ALP, GGT and total protein being positively correlated with Cd in urine. In conclusion, the workers in the tobacco leaf processing factory were found to be exposed to Cd compared to the general population. The increase in the urinary enzymes and proteins suggests that an exposure to Cd affects kidney functions even below the toxic limits generally accepted.

Carnitine metabolism in diabetes mellitus.

In diabetes mellitus (DM), increased fatty acids have negative effects on pancreatic beta-cell functions, in addition to enhanced mitochondrial transportation of fatty acids related to decreased insulin levels. The aim of this study was to evaluate lipid metabolism in children with DM by measuring plasma fatty acids and carnitine fractions to reveal relationships between carnitine status and increased fatty acid oxidation. Increased plasma fatty acids (except for arachidonic acid, there were no significant differences in the ratio of each specific fatty acid to total fatty acids), lipoprotein (a), acyl carnitine levels and urinary total and free acyl carnitine excretion, and decreased plasma free carnitine levels, were found in children with DM. There were no correlations between the duration of DM or HbA1c and study parameters. It is recommended that plasma free carnitine determinations should be made even if the patient has good metabolic control.