Adipocyte "Fatty Acid Binding Protein" Gene Polymorphisms (rs1054135, rs16909196 and rs16909187) in Jordanians with Obesity and Type 2 Diabetes Mellitus.

Background: Obesity, type 2 diabetes mellitus (T2DM), and dyslipidemia may result from the interactions of genetic and environmental factors. There are controversial reports concerning the association of polymorphisms (rs1054135, rs16909196 and rs16909187) in the gene of adipocyte fatty acid binding protein (FABP4) with obesity and T2DM. Therefore, we designed this study to determine the association of these polymorphisms with obesity, T2DM, and dyslipidemia among Jordanian subjects. Methods: The study was approved by the National Center for Diabetes, Endocrinology, and Genetics (NCDEG) Institutional Review Board (IRB). A total of 397 subjects were enrolled in the study and divided into four groups as described in materials and methods section. The fatty acid binding protein 4 (FABP4) gene containing (rs1054135, rs16909196 and rs16909187) single nucleotide polymorphisms (SNP) was amplified by polymerase chain reaction (PCR) followed by Sanger DNA sequencing of the PCR product. Results: None of the three SNPs were associated with T2DM (p > 0.05). The rs16909187 and rs16909196 were significantly associated with obesity. The wild type (CC) of rs16909187 was significantly higher among the overweight and obese group compared with normal weight controls (OD = 2.17, 95% CI = 1.18 - 3.96, p =0.01). The wild type of rs16909196 (AA) was significantly higher among the overweight and obese group compared to controls, (OD = 2.26, 95% CI = 1.24 - 4.14, p = 0.01). These results may indicate that the wild-type may be a risk factor for obesity.Only the rs1054135 SNP was significantly associated with increased low density lipoprotein (LDL) levels in the overweight and obese group compared with the controls (p = 0.03). Conclusions: The wild-type genotypes of rs16909196 and rs16909187 may be risk factors for obesity but not T2DM. None of the three SNPs was associated with T2DM.

Pharmacokinetics of vancomycin in neonates admitted to the neonatology unit at the Jordan University Hospital.

BACKGROUND: Vancomycin is used in neonatal intensive care units to treat nosocomial infections caused by methicillin-resistant Staphylococcus aureus (MRSA) or coagulase-negative staphylococci. Current dosing regimens are not adequate, producing trough concentrations below the target range. PATIENTS AND METHODS: 151 neonates who received vancomycin and had vancomycin peak and trough concentration measurements as part of regular therapeutic drug monitoring were utilized. The patients were divided into three groups according to gestational age; less than 28 weeks, 28 to less than 34 weeks and 34 weeks to term. Neonates were given vancomycin doses as per guidelines. The pharmacokinetic parameters, elimination rate constant (K), elimination half-life (t1/2), volume of distribution (VD) and clearance (CL), were calculated. RESULTS: Peak vancomycin concentrations ranged from 9.4 to 52.8 mg/l, while troughs ranged from 0.0 to 16.6 mg/l. One third of the trough concentrations were below 5 mg/l and are thus, subtherapeutic. There were no statistically significant differences between the three groups in regard to elimination rate constant and half-life. However, there was a statistically significant difference in volume of distribution between the three groups (p < 0.05), and in clearance between the first group and the other two (p < 0.05), using Kruskal-Wallis statistical test. CONCLUSIONS: The empirical dosing method used was inadequate in one third of patients. Individualization of vancomycin dosing in the neonatal critical care unit at The Jordan University Hospital seems necessary.

N-Acetyltransferase-2 (NAT2) genotype frequency among Jordanian volunteers.

OBJECTIVE: To determine the frequency of major NAT2 alleles and genotypes among the Jordanian population. METHODS: DNA samples from 150 healthy Jordanian volunteers were analyzed by polymerase chain reaction followed by restriction fragment length polymorphism assays (PCR-RFLP) to determine the frequency of four major alleles: NAT2*4, NAT2*5, NAT2*6 and NAT2*7. RESULTS: The frequency (95% confidence interval) of NAT2*4, NAT2*5, NAT2*6 and NAT2*7 alleles was 0.247 (0.198 - 0.296), 0.373 (0.318 - 0.428), 0.347. 0.293 - 0.401) and 0.033 (0.013 - 0.053), respectively. The most prevalent genotypes are those which encode slow acetylation phenotype. Around 58.7%, 33.3% and 8% of volunteers carried the slow, the intermediate and the fast-encoding genotypes, respectively. CONCLUSION: NAT2 genotype profile among Jordanians is similar to Caucasians. However, NAT2*6 allele is slightly high in comparison with Arab Middle Eastern populations.

Glucuronidation of 6 alpha-hydroxy bile acids by human liver microsomes.

The glucuronidation of 6-hydroxylated bile acids by human liver microsomes has been studied in vitro; for comparison, several major bile acids lacking a 6-hydroxyl group were also investigated. Glucuronidation rates for 6 alpha-hydroxylated bile acids were 10-20 times higher than those of substrates lacking a hydroxyl group in position 6. The highest rates measured were for hyodeoxy- and hyocholic acids, and kinetic analyses were carried out using these substrates. Rigorous product identification by high-field proton nuclear magnetic resonance and by electron impact mass spectrometry of methyl ester/peracetate derivatives revealed that 6-O-beta-D-glucuronides were the exclusive products formed in these enzymatic reactions. These results, together with literature data, indicate that 6 alpha-hydroxylation followed by 6-O-glucuronidation constitutes an alternative route of excretion of toxic hydrophobic bile acids.

Compliance with good practice in prescription writing at outpatient clinics in Saudi Arabia.

A sample of prescription orders received from outpatient departments by a hospital pharmacy in Asir, Saudi Arabia, were analysed over 1 year for the essential elements of prescriptions. The prescriber's name, address and signature were on 83.3%, 9.6% and 81.9% of prescriptions respecti-vely. The patient's name, age and sex were on 94.6%, 77.3% and 51.3%. No prescription contained the patient's address and weight. Generic drug names were used in only 15.1% and strength of medication and dose units were included in 26.6% and 55.6% of prescriptions. Most prescriptions (94.0%) had no quantity indicated and had only partial instructions for patient use (90.7%); the diagnosis was included in about two-thirds. The prescriber's handwriting was illegible in 64.3% of prescriptions. Measures to improve the situation are suggested.

S-mephenytoin hydroxylation phenotypes in a Jordanian population.

We tested the ability of 194 unrelated, healthy Jordanian volunteers to metabolize S-mephenytoin. Mephenytoin (100 mg) was coadministered with debrisoquin (10 mg) orally and urine was collected for 8 hours. Mephenytoin metabolism was tested according to three measures: the amount of 4-hydroxymephenytoin, the S/R enantiomeric ratio, and the presence of a polar, acid-labile metabolite in urine collected for 8 hours after the dose. The S/R ratio and the presence of the acid-labile metabolite were determined in the urine of 16 patients who had low amounts of 4-hydroxymephenytoin (log hydroxylation index > or = 1). On examination of these three parameters of oxidation status, nine subjects were found to be poor metabolizers of mephenytoin by all three parameters. Thus 4.6% (95% confidence interval of 1.6% to 7.6%) of Jordanian subjects studied were poor metabolizers of mephenytoin. According to the Hardy-Weinberg Law, the frequency of the recessive autosomal gene controlling the poor metabolizer status of mephenytoin was predicted to be 0.215% (95% confidence interval of 0.146% to 0.283%). These results are on the same order of magnitude as those determined in European white populations and constitute the first report in Arab populations.

Evaluation of therapeutic drug monitoring of antiepileptic drugs.

Results of the therapeutic drug monitoring (TDM) of the concentration of antiepileptic drugs (AEDs), performed at Aseer Central Hospital over a 1-year period were evaluated. Most of the requests were for phenytoin (PHT) determination followed by carbamazepine (CBZ), valproic acid (VPA) and phenobarbital (PB). Serum concentrations of PB, CBZ, VPA and PHT within the presumed therapeutic range constituted 87%, 73%, 45% and 33%, respectively. Valproic acid exhibited age differences in the proportion of concentrations below the presumed therapeutic range, such that subtherapeutic concentrations increased while therapeutic concentrations decreased with age. When the requests were related to particular patients, 71% of patients were on monotherapy with PHT as the most commonly used single drug and PB the least. Patients using 2 AEDs were found to constitute 23.5% of all patients with "PHT + CBZ" and "CBZ + VPA" being the most commonly prescribed 2 drugs combination. The frequency of concentrations within the therapeutic ranges decreased when 1 or 2 more drugs were used with either PHT or VPA. In addition, combination with PHT was associated with a reduction in mean CBZ concentration, while combination with CBZ was associated with a reduction in mean VPA concentration.

Comparative pharmacokinetics of 2 brands of cefuroxime following a single intramuscular injection.

The pharmacokinetic parameters (AUC0-8h, AUC0-infinity, Cmax, tmax, t1/2, Ke) of cefuroxime following a single intramuscular injection of 750 mg of a test product (Maxil, Hikma Pharmaceuticals, Amman, Jordan) were compared to those of a reference product (Zinacef, Glaxo, UK). The 2 products were administered according to a randomized 2-way crossover design to 26 healthy male volunteers. Cefuroxime plasma concentrations were determined using a rapid, sensitive and precise HPLC method with UV detection at 280 nm. The means of the ratios AUC0-infinity Maxil/AUC0-infinity Zinacef and Cmax Maxil/Cmax Zinacef were close to 1 and their 90% confidence intervals included 1. The mean pharmacokinetic parameters of the 2 products were not different and their 90% confidence intervals overlapped. The 2 products were not statistically different with respect to both rate and extent of absorption as demonstrated by statistical analysis on Cmax, tmax, AUC0-8h and AUC0-infinity. The 2 products were also similar regarding t1/2 and Ke. The ANOVA analysis showed no differences in the pharmacokinetic parameters of the 2 products in relation to treatment, sequence of product administration or the interaction term. Pharmacokinetic parameters of cefuroxime were comparable to reported values. We conclude that the 2 products of cefuroxime (Maxil and Zinacef) are bioequivalent.

Comparative pharmacokinetics of two brands of atenolol following a single oral administration.

The pharmacokinetic parameters (Cmax, Tmax, t1/2, AUC0-30h, AUC0-infinity) of following a single oral administration of 100 mg of a test product (Tenolol, The United Pharmaceutical Manufacturing Company, Amman, Jordan) were compared to those of a reference product (Tenormin, ICI Pharmaceuticals). The 2 products were administered according to a randomized 2-way crossover design to 24 healthy male volunteers. After drug administration, serial blood samples were collected over a period of 30 hours. Atenolol plasma concentrations were measured using an HPLC technique with fluorometric detection at an excitation and an emission wavelengths of 222 nm and 300 nm, respectively. The parametric 90% confidence intervals of the mean value of the ratio (Tenolol/Tenormin) of pharmacokinetic parameters were 0.90-1.12, 0.92-1.12, 0.88-1.14, and 0.91-1.09 for AUC0-30h, AUC0-infinity, Cmax, and t1/2. In each case values were within the acceptable bioequivalence range of 0.8-1.25. Point estimates of these parameters were 1.01, 1.02, 1.01, and 1.0. The parametric point estimate of the mean difference of Tmax between the 2 formulations (Tenolol-Tenormin) was 0.19 hours with a 90% confidence interval of -0.47-0.84, which overlaps with the stipulated bioequivalence range of +/- 0.64. Thus, the 2 products could be considered bioequivalent regarding rate of absorption (Cmax and Tmax), extent of absorption (Cmax and AUC), and elimination (t1/2).

Comparative pharmacokinetics of two nifedipine products in capsule form following single oral administration in healthy volunteers.

The pharmacokinetic parameters (AUC, Cmax, Tmax, and t1/2) of nifedipine following single oral administration of a 10 mg capsule of test product were compared to those of the same amount of a reference product. The two products in capsule form were administered according to a randomized two-way crossover design in 22 healthy male volunteers. Nifedipine plasma concentrations were determined using a rapid, sensitive and precise high performance liquid chromatography (HPLC) method with ultraviolet (UV) detection at 235 nm. The parametric 90% confidence intervals of the mean value of the ratio [Myogard (test product) /Adalat (reference product)] for pharmacokinetic parameters were 0.90-1.08, 0.80-1.07, and 0.93-1.12 for AUC0-->infinity, Cmax and t1/2, respectively. In each case, values were within the acceptable bioequivalence range of 0.8-1.25. Distribution free point estimate for the difference in expected medians of Tmax of the two products (Myogard-Adalat) was 0.00 h with a 90% confidence interval of 0.00-0.13 which is greater than the accepted bioequivalence of +/- 0.12. The kinetic parameters were comparable to those reported for nifedipine, and no statistically significant differences were found in any of them when comparing the two products by analysis of variance (ANOVA) on log-transformed data. Thus, the two products could be considered bioequivalent regarding absorption rate (Cmax and Tmax), extent of absorption (Cmax and AUC) and elimination (t1/2).

Glucuronidation of 7-hydroxy-4-methylcoumarin by human liver microsomes. Inhibition by certain drugs.

Glucuronidation of 7-hydroxy-4-methylcoumarin (7-OH-4-MC) was comparable in three of the four human liver samples studied. In liver sample IV the glucuronidation rate of 7-OH-4-MC was almost 40% of that in the other samples. That might be due to interindividual variation in the capacity to glucuronidate. Glucuronidation rates were not reduced in the one human liver sample which displayed mild centrilobular cholestasis on histopathology. Furosemide (FR), salicylic acid (SA), lorazepam (Z) and menthol (MT) inhibited the glucuronidation of 7-OH-4-MC to varying degrees (30-90%). Paracetamol (PA) and 5-hydroxytryptamine (HT) were not inhibitors of the glucuronidation of 7-OH-4-MC. These findings suggest the presence of other UDP-glucuronosyltransferase (UDPGT) isozymes in the human liver microsomes which are responsible for the glucuronidation of PA and HT than those which glucuronidate 7-OH-4-MC.

Dextromethorphan metabolism in Jordanians: dissociation of dextromethorphan O-demethylation from debrisoquine 4-hydroxylation.

The concentrations of dextromethorphan (DM) and its metabolites dextrorphan (DRP), 3-methoxymorphinan (MM) and 3-hydroxymorphinan (HM) were measured in 8 h urine samples from 266 unrelated healthy Jordanian subjects following oral administration of 30 mg dextromethorphan hydrobromide and using a rapid, sensitive and precise HPLC method with fluorometric detection. The frequency of the 'poor' metabolizer status of DM-O-demethylation as judged by log DM/DRP was found to be 6.8% with a 95% confidence interval of 3.8-9.8%. There was a strong correlation between log DM/DRP and log total non-O-demethylated compounds (NODM)/total O-demethylated metabolites (ODM) metabolic ratios (r = 0.96, P < 0.01). However, one subject with log DM/DRP of 0.05 that classifies him as a poor metabolizer was found to have a log NODM/ODM of -0.73 which is in the range of extensive metabolizer status suggesting the presence of another cytochrome P450 isoenzyme involved in dextromethorphan O-demethylation. Dextromethorphan N-demethylation to 3-methoxymorphinan was detected in 55.3% of individuals. Furthermore, a dissociation between dextromethorphan O-demethylation and debrisoquine (D) 4-hydroxylation has been observed. Among the 116 subjects phenotyped with both dextromethorphan and debrisoquine, 7 were poor metabolizers of both, three were poor metabolizers of debrisoquine and extensive metabolizers of dextromethorphan whilst 4 were poor metabolizers of dextromethorphan and extensive metabolizers of debrisoquine, one of whom was reclassified as an extensive metabolizer of dextromethorphan using log NODM/ODM to characterize dextromethorphan metabolizer status.

Isolation and purification of two human liver UDP-glucuronosyltransferases.

Two UDP-glucuronosyltransferases (EC 2.4.1.17) were purified from human liver microsomes. Human liver microsomes were solubilized with Emulgen 911 and the UDP-glucuronosyltransferases were separated and purified by chromatofocusing and UDP-hexanolamine Sepharose 4B affinity chromatography. One isoenzyme eluted with an apparent pl of 7.4, displayed a subunit molecular weight of 53,000 after sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and catalyzed the glucuronidation of p-nitrophenol, 4-methylumbelliferone, alpha-naphthylamine, and estriol, but not that of 4-aminobiphenyl. A second isoenzyme eluted with an apparent pl of 6.2, displayed a subunit molecular weight of 54,000 after SDS-PAGE, and catalyzed the glucuronidation of p-nitrophenol, 4-methylumbelliferone, alpha-naphthylamine, and 4-aminobiphenyl, but not that of estriol. Neither of the purified human liver UDP-glucuronosyltransferases employed estrone, beta-estradiol, testosterone, androsterone, or 5 alpha-androstane-3 alpha,17 beta-diol as substrate. These enzymes displayed apparent Km values in the same order of magnitude for a given substrate. In general, high concentrations of phosphatidylcholine were required for reconstitution of maximal glucuronidation activity. This report documents the existence of multiple UDP-glucuronosyltransferases in human liver.

Characterization of antibodies to a rabbit hepatic UDP-glucuronosyltransferase and the identification of an immunologically similar enzyme in human liver.

An antibody to a UDP-glucuronosyltransferase (UDPGT) isoenzyme which catalyzes the glucuronidation of p-nitrophenol (PNP) in rabbit liver was raised in sheep and used to identify immunologically similar UDPGTs in rabbit and human livers. Immunoblotting experiments showed that the antisera specifically recognized PNP UDPGT but not estrone UDPGT purified from rabbit liver. Sheep anti-rabbit liver PNP UDPGT IgG immunoprecipitated PNP, 1-naphthol, and 4-methylumbelliferone glucuronidation activities in rabbit and human liver microsomal preparations. In rabbit liver microsomes the antibody did not immunoprecipitate estrone or estradiol glucuronidation activities. In human liver microsomes, 4-aminobiphenyl but not estriol glucuronidation activities were immunoprecipitated, suggesting that the antibody recognizes a specific UDPGT (pI 6.2) in human liver microsomes.

Glucuronidation of monohydroxylated short chain bile acids by human liver microsomes and purified human liver UDP-glucuronosyltransferases.

The glucuronidation of monohydroxylated bile acids and their analogs with a shortened side chain (short-chain bile acids) by human liver microsomes and by two UDP-glucuronosyltransferases purified therefrom has been studied in vitro. In microsomes, all 18 substrates tested underwent glucuronidation; the rate of reaction and the site of attachment of glucuronic acid (hydroxyl group in position 3, side chain carboxyl group, or various ratios of both products) were strongly dependent on the length of the side chain, the configuration of the 3-hydroxyl group, and the configuration of the A/B ring junction (5 alpha-H/5 beta-H). Two UDP-glucuronosyltransferases (UDPGTs) purified from human microsomes, designated "pl 7.4" and "pl 6.2" according to their behavior in chromatofocusing, accounted for the formation of hydroxyl-linked glucuronides of a different pair of bile acids each. The pl 7.4 human liver UDPGT catalyzed the glucuronidation of C20 and C22 with the 3 alpha-OH, 5 beta-H configuration, while the pl 6.2 human liver UDPGT catalyzed the glucuronidation of either 3-OH epimer of C21 and C24 acids with the 5 alpha-H configuration. The enzymes displayed a relatively high selectivity in that they did not accept any of the remaining 14 bile acids as substrates; none of the enzymes led to the formation of a carboxyl-linked glucuronide. In addition, purified human liver UDPGT did not catalyze the glucuronidation of cholate, deoxycholate, chenodeoxycholate, ursodeoxycholate, lithocholate, hyocholate, hyodeoxycholate, bilirubin, morphine, or 4-hydroxybiphenyl. The above results suggest that several bile-acid UDP-glucuronosyltransferases of high specificity exist in human liver.

N-acetylation phenotyping using dapsone in a Jordanian population.

1. The N-acetylation of dapsone (DDS) was studied in 160 unrelated healthy Jordanian volunteers. 2. The frequency of slow acetylators determined using the plasma monoacetyldapsone (MADDS) to DDS ratio (MADDS/DDS), was 67.5% with a 95% confidence interval of 59 to 76%. Slow acetylators had an acetylation ratio of less than 0.42. 3. Applying the Hardy-Weinberg Law, the frequency of the recessive allele controlling slow acetylation was found to be 0.82 +/- 0.02. 4. The frequency distribution histogram of the plasma MADDS/DDS ratio showed an apparent trimodal pattern. The number of homozygous (n = 16) and heterozygous (n = 36) rapid acetylators derived from the observed data did not agree with those predicted for the respective rapid acetylators (n = 5 and n = 47) according to the Hardy-Weinberg Law. The suggested antimode used to discriminate the two groups was 0.82. 5. The mean plasma concentration of MADDS and the mean plasma acetylation ratio were about three times lower in slow than in rapid acetylators. However, there was no difference in mean plasma DDS concentration between slow and rapid acetylators. 6. There was a significant correlation (r = 0.853, P less than 0.001) between plasma MADDS concentration and the acetylation ratio. For DDS such a correlation was absent (r = 0.059, P = 0.23).

Simultaneous high performance liquid chromatographic determination of dapsone and monoacetyldapsone in human plasma and urine.

A rapid, specific and a one-stage protein precipitation method for simultaneous estimation of dapsone (DDS) and monoacetyldapsone (MAD) concentration in plasma and urine using high performance liquid chromatography (HPLC) is described. The applicability of the method for monitoring DDS and MAD blood levels in two different acetylator phenotype volunteers following the administration of 100-mg oral dose of DDS was shown. Cumulative urinary excretion of DDS and MAD were studied in the same volunteers.

Gastrointestinal dialysis of digoxin using cholestyramine.

The pharmacokinetic parameters of digoxin given intravenously (0.075 mg/kg) alone and following treatment with oral cholestyramine (8 gm in 50 ml water) were studied in rabbits. Pretreatment with cholestyramine produced a significant decrease in the serum concentration of digoxin and significantly enhances its systemic clearance as indicated by a statistically significant decrease in the area under the concentration-time curve (AUC), half time of elimination (t 1/2), and mean residence time (MRT). These findings indicate that the idea of gastrointestinal dialysis, known with activated charcoal, could be extended to ion-exchange resins that could be a potentially useful adjunctive measure in the management of drug overdose especially with commonly used drugs with a low therapeutic index like digoxin.

Dextromethorphan O-demethylation polymorphism in Jordanians.

The O-demethylation of dextromethorphan (DMT) to dextrorphan (DRP) was studied in 241 unrelated, healthy Jordanian volunteers (171 males, 70 females). Urine was collected for 8 h following a single oral dose of DMT bromhydrate 30 mg. A thin-layer chromatographic (TLC) technique was used to identify the metaboliser phenotype. The frequency of the poor metaboliser phenotype was found to be 2.9% (approximate 95% confidence interval 0.8-5.0%). Applying the Hardy-Weinberg Law, the frequency of the recessive autosomal gene controlling poor metabolism was 0.17 (95% confidence interval 0.108-0.232).