Gas chromatography-mass spectrometric analysis of clenbuterol from urine.

Clenbuterol which is mostly used as an anabolic agent. It is also used for treatment of asthma. Clenbuterol was analysed from urine by using gas chromatography-mass spectrometry. GC-MS parameters were determined. Timolol was used as an internal standard. Extraction and derivatisation procedure of clenbuterol from urine were developed. Clenbuterol was extracted by using diethylether/ter-butanol (4:1; v:v) and pH 12 K2CO3/KHCO3 (3:2; w:w) buffer. MSTFA/NH4I (1 ml/10 mg) mixture was used for derivatization of clenbuterol. Selected ions of clenbuterol-bis-TMS were m/z: 405, 337, 336, 335, 300, and 227. Extraction yield and minimum detection limit of clenbuterol from urine were identified. Extraction yield was 94.30% and minimum detection was found 0.02 ng ml(-1) urine. It has been concluded that the GC-MS method is sensitive, accurate, precise, and reproducible for analysing of clenbuterol from urine.

Optimization of mobile phase in the separation of beta-blockers by HPLC.

Beta-blockers are generally determined using high-performance liquid chromatography (HPLC). Previous HPLC separations of beta-blockers have often required a mobile phase containing three components; acetonitrile or methanol to control the retention; buffer to control the ionic strength and pH of the mobile phase; ion-pairing reagent to provide adequate retention of beta-blockers or organic amines as masking agent to reduce peak tailing. Due to the complexity of the mobile phases employed, development of these assays can be a laborious process. Additionally, alkyl sulphonates and organic amines dramatically reduces the life-time reduction of silica based C18 columns. The results of this study demonstrated that the addition of tested alkyl sulphonates and organic amines is not essential for an adequate separation of beta-blockers. In this study, we developed a simple HPLC method for the simultaneous separation of model beta-blockers, atenolol, practolol, metoprolol, oxprenolol and propranolol. Atenolol, practolol, metoprolol, oxprenolol and propranolol adequately separated with high peak symmetries using a mobile phase consisted of methanol/acetonitrile/phosphate buffer (10 mM, pH 3.0) (15:15:70, v/v/v). By altering only the fraction of methanol with respect to acetonitrile, method development becomes a more efficient separation. Furthermore, atenolol, practolol, metoprolol, oxprenolol and propranolol can be detected up to 0.25, 5, 10, 50 and 10 ng ml(-1). In this publication, we present the simultaneous separation of beta-blockers having a wide range of polarity. It is proposed that this new mobile phase, consisting only acetonitrile, methanol and phosphate buffer can be used for the analysis of the several beta-blockers presently in doping control analysis as well as others.

The effect of interferon and desferrioxamine on serum ferritin and hepatic iron concentrations in chronic hepatitis B.

BACKGROUND/AIMS: Recent reports indicate that an individual's iron status might affect the response rate achieved with Interferon therapy for the treatment of chronic viral hepatitis. METHODOLOGY: Forty individuals, 29 men and 11 women, with chronic viral hepatitis B, who had elevated serum ferritin levels, were randomized to receive either Interferon (IFN) 5 MU TIW SQ for 6 months alone (n=21) or Interferon in combination with repetitive cycles of desferrioxamine infused at a dose of 80 mg/kg per cycle (n=19) over 3 consecutive days in an effort to reduce their metabolically active iron pool during the course of IFN treatment. These cycles were continued until a serum ferritin level of less than 250 ng/ml (normal values <220 ng/ml) was achieved. Additionally, all desferrioxamine treated subjects were placed on a low iron containing diet. An interferon response was defined as normalization of the serum ALT and seroconversion from eAg positive to eAb positive. All other responses were defined as failures. RESULTS: The mean ages of the subjects in the 2 groups were 39+/-6 and 38+/-5 years. The initial serum ALT levels were 150+/-27 and 151+/-13 IU/l. The hepatic iron concentrations were 916+/-29 and 896+/-15 microg/g/dry liver weight. The serum ferritin levels were 386+/-12 and 393+/-18 ng/ml. None of these values differed significantly between the 2 treatment groups. The desferrioxamine treated group consisted of 14 men and 5 women. This group experienced a reduction in their serum ferritin to a level of 237+/-13 ng/ml as a result of the desferrioxamine treatment (p<0.05). Additionally, a reduction in their hepatic iron concentration, to a level 766+/-29 microg/g/dry liver weight, occurred with treatment (p<0.05). Twelve of the 19 (63%) desferrioxamine-treated subjects and 8 of the 21 (38%) control subjects experienced a normalization of their serum ALT levels with treatment (p<0.05). Thirteen of 19 (68%) of the desferrioxamine-treated subjects but only 8 of 21 (38%) of the IFN alone treated group seroconverted to anti-e positive (p<0.05). Moreover, a greater improvement in the hepatic histologic score and rate of HBV-DNA loss occurred in the desferrioxamine-treated group. CONCLUSIONS: Based upon these data, it can be concluded that desferrioxamine infusion to achieve a normal serum ferritin level enhances the likelihood of an individual with chronic hepatitis B responding to IFN therapy. The precise mechanism responsible for this phenomenon is not clear, but would appear to be due to a reduction in the hepatic free iron pool as reflected by sequential changes in the serum ferritin and hepatic iron concentrations.

The determination of nandrolone and its metabolites in urine by gas chromatography-mass spectrometry.

The determination of nandrolone and its major metabolites in urine of a healthy volunteer is typically performed by fused-silica capillary column gas chromatography with electron impact quadropole mass spectrometry. Two well-known urinary metabolites of nandrolone, 19-norandrosterone and 19-norethiocholanolone, were isolated by XAD-2 adsorption from urine, eluted with methanol and separated into unconjugated and conjugated fractions. The conjugated fraction was hydrolyzed with beta-glucuronidase from Escherichia coli and the samples derivatized with MSTFA/ammonium iodide/dithioerythritol. Ion fragmentograms of the bis-trimethylsilyl derivatives of nandrolone and its metabolites displayed molecular ions of M+ = 420 and M(+) -15 = 405. Extraction yield and the minimum detection limit of nandrolone in urine were identified. Finally, excretion rates of nandrolone and its metabolites in urine were determined.

The use of deferoxamine infusions to enhance the response rate to interferon-alpha treatment of chronic viral hepatitis B.

An individual's iron status may affect the response rate achieved with the use of interferon (IFN) as therapy for chronic viral hepatitis. A total of 27 patients with chronic hepatitis B viral infection, who had elevated serum ferritin levels, were randomized to receive either IFN 5 MU, three times weekly by subcutaneous injection alone (n = 14) or in combination with cycles of deferoxamine at a dose od 80 mg kg-1 per cycle (n = 13) administered over 3 consecutive days, to reduce their iron and maintain a serum ferritin level less than 250 ng ml-1. All deferoxamine-treated patients were on a low iron-containing diet. An IFN response was defined as a normalization of the serum alanine aminotransferase (ALT) level and seroconversion from hepatitis B e antigen (HBeAg) positivity to hepatitis B e antibody (HBeAb) positivity. The deferoxamine-treated group experienced a reduction in their serum ferritin level to 226 +/- 73 ng ml-1 as a result of the deferoxamine treatment. Six of the 13 (46%) deferoxamine-treated patients and two of the 14 (14%) control patients normalized their ALT levels. Seven of the 13 (54%) deferoxamine but only 14% of the IFN-treated group seroconverted to HBeAb positivity. A greater rate of histological improvement and loss of hepatitis B virus (HBV) DNA was seen in the deferoxamine-treated group. Two of the deferoxamine-treated patients were treated only once, two were treated twice, seven were treated three times and two were treated four times to achieve a ferritin level below 250 ng ml-1. Based on these data, we conclude that deferoxamine infusion enhances the rate of response to IFN in subjects with chronic hepatitis B. The precise mechanism of this phenomenon is not clear.

Relationship between the serum alanine aminotransferase level at the end of interferon treatment and histologic changes in wild-type and precore mutant hepatitis B virus infections.

Unravelling the role of interferon (IFN) in the treatment of chronic hepatitis B compliance by many factors. Several mutant forms of hepatitis B virus (HBV) have recently been discovered; the most common of these is the precore mutant, characterized by hepatitis B e antigen (HBeAg) negativity and hepatitis B e antibody (HBeAb) positivity in an individual with an active HBV infection. The aim of this study was to compare the response rate to IFN therapy in patients with wild-type HBV infection and in individuals infected with the precore mutant. A second aim was to evaluate the role of an increased serum ferritin in terms of the IFN response rate in these two different types of HBV infection. IFN therapy was administered at a dose of 5 MU subcutaneously three times weekly for 6 months to 41 individuals with a chronic wild-type hepatitis B infection and 16 individuals with a precore mutant chronic HBV infection. An IFN response was defined as normalization of the serum alanine aminotransferase (ALT) level and an HBeAb to HBeAb seroconversion (in wild-type hepatitis infection), and a normalization of the serum ALT in individuals infected with a precore mutant infection. At entry, the two groups were matched for age, gender, serum ALT, serum iron, total iron binding capacity (TIBC), serum ferritin and liver histology. Forty-six per cent of the subjects with wild-type disease responded to IFN therapy. By contrast, only four of the 16 cases (25%) of the precore mutant cases responded (p < 0.05). Ferritin levels correlated well with the type of IFN response; as the serum ferritin level increased, the response rate to IFN declined. Hapatic infection caused by a precore HBV mutant is more resistant to IFN therapy than wild-type infection. The serum ferritin level appears to influence the type of IFN response achieved. Individuals with a serum ferritin level greater than 300 ng ml-1 failed to respond to IFN in 93% of the cases studied.

Determination of ceftriaxone in aqueous humor and serum samples of differential-pulse adsorptive stripping voltammetry.

Differential-pulse adsorptive stripping voltammetry was used to determine ceftriaxone in serum and aqueous humour samples. The method involved extraction of the ceftriaxone from serum samples with an Amberlite XAD-2 column followed by elution with methanol. The recovery was 97.6% with a relative standard deviation of 3.3% at a ceftriaxone concentration of 90.9 microg 1(-1). Peak currents of ceftriaxone were measured with a hanging mercury drop electrode at -0.78 V versus an Ag-AgCl reference electrode in pH 3.0 Britton-Robinson buffer. The calibration graph was linear from 0.02 to 1300 microg 1(-1). The method was applied to cataract cases and ceftriaxone levels were measured in aqueous humour and serum samples from patients who had received 1 or 2 g of ceftriaxone intravenously. Aqueous humour was added to the polarographic cell directly. The amounts of ceftriaxone in the aqueous humour and serum samples with respect to time were measured. The pharmacokinetic profiles for 1 and 2 g were compared.

Determination of aluminum levels in the kidney, liver, and brain of mice treated with aluminum hydroxide.

In the present study, aluminum (Al) accumulation has been examined after aluminum loading in mice. The kidney, liver, and brain aluminum levels for mice that had been treated orally with aluminum hydroxide for 105 d and for the control group were determined using graphite furnace atomic absorption spectrophotometry (GFAAS) following an acid digestion. Matrix modifier consisted of 2% Triton X-100 and 2% Mg (NO3)2. Al loaded mice showed a significant increase in tissue aluminum levels, relative to the control group.

Plasma chromium levels in small-for-gestational-age newborn infants.

Plasma chromium (Cr) levels were determined in 24 preterms and 18 full-term newborn infants. There was no statistically significant differences in plasma Cr levels between the preterm and full-term infants. Plasma Cr levels were similar in small-for-gestational-age infants and in infants with hypoglycemia compared with healthy infants.

Determination of ceftriaxone in biological material by differential-pulse adsorptive stripping voltammetry.

Differential-pulse adsorptive stripping voltammetry was used to determine sub-micromolar concentrations of ceftriaxone in plasma. A hanging mercury drop electrode was chosen as the working electrode. A simple clean-up procedure was developed in which ceftriaxone was extracted from blood plasma with the non-ionic resin Amberlite XAD-2 and eluted with methanol. The recovery from plasma was 97.6% using a 1.52 x 10(-4) M stock ceftriaxone solution. The method was applied to caesarean cases, and total ceftriaxone levels were measured in the maternal and umbilical cord blood. The amount of ceftriaxone transmitted to the baby on administration of the drug to the mother before the caesarean operation was found to be in the range 0.067-0.17%.

Differential pulse adsorptive stripping voltammetric determination of ceftriaxone at a static mercury dropping electrode.

Ceftriaxone is a member of the "third generation" of cephalosporin antibiotics which adsorbs strongly onto a mercury electrode. By using this phenomenon and by accumulating this compound at a static mercury dropping electrode prior to differential pulse voltammetric measurements, very high sensitivities can be readily achieved. The influence of several variables (including accumulation time, modulation amplitude, rest period, scan rate, and drop size) on the adsorptive stripping response has been evaluated. Peak currents were measured with a hanging mercury dropping electrode at -0.78 V versus an Ag/AgCl reference electrode in pH 3.0 Britton-Robinson buffer. The linear calibration range was 3.31 x 10(-11) to 2.17 x 10(-6) M.

In vitro effects of thyroxine on the mechanical properties of erythrocytes.

In vitro effects of thyroxine on erythrocyte deformability and mechanical fragility were observed. Deformability of erythrocytes was improved in a dose dependent manner by thyroxine. Mechanical hemolysis was found to be lower if thyroxine was included in erythrocyte suspensions at concentrations close to the physiological levels (10(-9)M). These changes might be related to the alterations of intracellular calcium concentration, as in the erythrocyte suspensions containing 10(-9)M thyroxine, intracellular calcium concentration was found to be 30 times lower than the control suspensions which did not contain thyroxine. Thyroxine also reduced the mechanical hemolysis ratio in calcium loaded cells. These observations suggest that thyroxine might play some role in the regulation of the mechanical properties of erythrocytes which might be mediated via the effects on calcium metabolism.

Serum calcium levels in various lung diseases.

Calcium levels were determined in sera of patients suffering from various lung diseases. Healthy volunteers served as controls. Significant differences were found between the serum calcium levels of patients with active lung tuberculosis and those of controls (P less than 0.01). After treatment, serum calcium levels decrease to normal values in these patients. It was also found that there were significant differences in serum calcium levels of patients with primary lung carcinoma (P less than 0.01) and of patients with metastatic lung carcinoma as compared to controls (P less than 0.01). On the other hand, normal serum calcium levels were found in patients with pulmonary diseases with or without an infection. In conclusion, it seems likely that a combination of mechanisms plays a role in the pathogenesis of hypercalcaemia in pulmonary tuberculosis and primary and metastatic lung carcinoma.

The quantitative investigation of infrared laser effects on the levels of copper and zinc in various tissues.

In order to study the effects of infrared laser irradiation on biological systems, the levels of copper and zinc in various irradiated tissues have been investigated. The zinc level in the muscle of the irradiated group increased (P less than 0.001) compared with the control group while in submandibular glands its level decreased (P less than 0.001). The copper levels in these two types of tissues decreased significantly (P less than 0.001). In irradiated skin, both copper and zinc levels increased compared with the control group (P less than 0.005). Quantitative determination of the trace elements copper and zinc indicated the necessity for more detailed studies of the effect of infrared laser irradiation at the cellular level.

Determination of methotrexate in human blood plasma by adsorptive stripping voltammetry.

The use of adsorptive stripping voltammetry to measure sub-micromolar concentrations of methotrexate in plasma has been investigated. A simple clean-up procedure has been developed in which methotrexate is extracted from blood plasma with Amberlite XAD-2, which is a non-ionic resin, and eluted with methanol. Recovery for plasma analysis was 80%.

Electroanalytical determination of vinca alkaloids used in cancer chemotherapy.

A differential pulse polarographic method has been developed for determination of the antineoplastic agents vincristine and vinblastine at ng ml level, in biological fluids such as plasma and urine. The vincristine and vinblastine are extracted from urine with Amberlite XAD-2. Linear calibration plots are obtained for both over the concentration range 0.005-5 mug ml . The relative standard deviations found were 1.7% for analysis of the pure drugs, 7.3% for urine and 8.6% for plasma.

Polarographic determination of antihistamines by complexation with Cd(II).

the polarographic behavior of the complexes formed by Cd(II) ion with ethanolamine derivative antihistamines such as diphenhydramine hydrochloride, dimenhydrinate, and chlorphenoxamine hydrochloride was studied. Antihistamines form spontaneous complexes with Cd(II) ion in the presence of KNO3. In addition, pH 8.00 borate buffer was added to increase the differential pulse polarogram peak height, and tetraalkyl ammonium salts were added to increase the linear range. The method of determination developed has been applied to commercial tablet, capsule, elixir, and injection forms of ethanolamine derivative drugs and has been compared with official methods.