An isocratic high performance liquid chromatography (HPLC) assay for moxifloxacin, a new 8-methoxyquinolone

OBJECTIVE: Studies have shown memory deficits among combat veterans with posttraumatic stress disorder (PTSD); however, high rates of comorbid conditions, including alcoholism, make it difficult to definitively associate these findings with the PTSD diagnosis. In this study the authors examined memory functioning among rape survivors without alcoholism or substance abuse but with PTSD. METHOD: Rape victims with (N = 15) and without (N = 16) PTSD were compared to age- and education-matched nontraumatized comparison subjects (N = 16) on measures of learning and memory. RESULTS: The subjects with PTSD performed significantly worse than the other groups on delayed free recall. The deficits were ameliorated by cueing and recognition testing. CONCLUSIONS: Recall deficits in noncombat PTSD patients strengthen the theory that memory deficits are associated with the PTSD diagnosis. The deficits were mild and were not attributable to comorbid depression, anxiety, or substance abuse.     

Comparison of stability-indicating assay methods: ultraviolet, high pressure liquid chromatography and acid dye

Three techniques--ultraviolet, high pressure liquid chromatography (HPLC) and acid dye--were compared as stability-indicating assay methods. Investigations on six drugs (codeine phosphate, ephedrine hydrochloride, morphine sulfate, procaine hydrochloride, pyrilamine maleate and thiamine hydrochloride) indicate that HPLC is the most reliable method.     

Overexpression, purification, and characterization of the cloned metallo-beta-lactamase L1 from Stenotrophomonas maltophilia

OBJECTIVES: The purpose of this study was to assess whether the presence or absence of myocardial viability during dobutamine echocardiography (DE) predicts survival in patients with coronary artery disease (CAD) and severe left ventricular (LV) dysfunction. BACKGROUND: In patients with CAD, the presence of myocardial viability during DE identifies viable myocardium and predicts recovery of LV systolic function after revascularization. However, there is little data on the relation between myocardial viability and clinical outcome in patients with CAD and severe LV dysfunction. METHODS: We studied 318 patients with CAD and a LV ejection fraction (EF) < or =35% who underwent DE and were followed for 18+/-10 months. Patients were classified into four groups. Group I (n=85) consisted of patients who had evidence of myocardial viability and subsequently underwent revascularization. Group II (n=119) consisted of patients with myocardial viability who did not undergo revascularization. Group III (n=30) consisted of patients who did not have myocardial viability and underwent revascularization. Finally, group IV (n=84) patients lacked myocardial viability and did not undergo revascularization. RESULTS: The four groups had similar baseline characteristics and rest LVEF. During follow-up there were 51 deaths (16%). The mortality rate was 6% in group I, 20% in group II, 17% in group III and 20% in group TV (p=0.01, group I vs. other groups). CONCLUSIONS: In patients with CAD and severe LV dysfunction who demonstrated myocardial viability during DE, revascularization improved survival compared with medical therapy.     

Reverse-phase capillary high performance liquid chromatography/high performance electrospray ionization mass spectrometry: an essential tool for the characterization of complex glycoprotein digests

The B-domain of recombinant human Factor VIII comprises 909 amino acids and is extensively N- and O-glycosylated, in that at least 20 different sites are occupied by numerous carbohydrate structures. This domain was incubated with trypsin and subjected to liquid chromatography electrospray ionization mass spectrometry analysis, using an electrospray orthogonal acceleration time-of-flight mass spectrometer as the detector for a capillary reversed phase HPLC separation of the digest. The inherent high mass resolution afforded by this instrument provides both ion charge state determination and high accuracy mass measurement that are of significant advantage in defining such highly complex mixtures.     

Purification and characterization of N-acetylglucosamine 6-phosphate deacetylase with activity against N-acetylglucosamine from Vibrio cholerae non-O1

An enzyme that deacetylates N-acetylglucosamine to glucosamine from Vibrio cholerae non-O1 was purified to homogeneity by sequential procedures. The native enzyme had a molecular mass of 190,000 Da and was predicted to be composed of four identical subunits with molecular masses of 45,000 Da. The purified enzyme hydrolyzed N-acetylglucosamine, N-acetylglucosamine 6-phosphate, and N-acetylglucosamine 6-sulfate, but not chitin oligosaccharides, and N-acetylgalactosamine. The deacetylase activity was completely abolished by N-ethylmaleimide, p-chloromercuribenzoate, EDTA, and Cu2+. On the other hand, the activity was activated by Co2+. The amino-terminal amino acids of the purified enzyme were sequenced. Among the 22 N-terminal amino acid residues, 12 residues of Vibrio deacetylase were identical with that of Escherichia coli GlcNAc 6-phosphate deacetylase.     

Isolation and molecular characterization of the Arabidopsis TPS1 gene, encoding trehalose-6-phosphate synthase

An Arabidopsis thaliana cDNA clone, AtTPS1, that encodes a trehalose-6-phosphate synthase was isolated. The identity of this protein is supported by both structural and functional evidence. On one hand, the predicted sequence of the protein encoded by AtTPS1 showed a high degree of similarity with trehalose-6-phosphate synthases of different organisms. On the other hand, expression of the AtTPS1 cDNA in the yeast tps1 mutant restored its ability to synthesize trehalose and suppressed its growth defect related to the lack of trehalose-6-phosphate. Genomic organization and expression analyses suggest that AtTPS1 is a single-copy gene and is expressed constitutively at very low levels.     

Quantitative analysis of ceramide molecular species by high performance liquid chromatography

BACKGROUND: Hepatocyte growth factor/scatter factor (HGF/SF) is a potent mitogen for various neoplastic cells, including neoplastic bronchial epithelia. METHODS: Immunoreactive hepatocyte growth factor/scatter factor (HGF/SF) was measured in extracts prepared from 129 nonsmall cell lung carcinoma (NSCLC) specimens, using an enzyme-linked immunosorbent assay. These specimens represented 5 cases of solitary/localized bronchioloalveolar cell carcinoma (BAC), 4 cases of diffuse/infiltrative BAC, 90 cases of non-BAC adenocarcinoma, 25 cases of squamous cell carcinoma, and 5 cases of large cell carcinoma. RESULTS: The mean concentration of immunoreactive HGF/SF was more than 19-fold higher in tissue extracts from diffuse-type BAG (265.0 +/- 110.2 ng/100 mg protein) than in those from solitary-type BAC (13.9 +/- 15.9, P < 0.005), non-BAC adenocarcinoma (13.8 +/- 14.9, P < 0.001), squamous cell carcinoma (13.2 +/- 14.4, P < 0.001), or large cell carcinoma (11.2 +/- 6.5, P < 0.005). When immunohistochemical staining for HGF/SF was performed, intense HGF/SF staining was uniformly observed in diffuse-type BAC tumor cells, but not in solitary-type BAC. CONCLUSIONS: Although BAC is included as a subtype of adenocarcinoma in the World Health Organization classification, diffuse-type BAC should be considered a distinct biologic entity, at least in terms of HGF/SF expression, from solitary-type BAC or non-BAC adenocarcinoma. In addition, the solitary and diffuse forms of BAC are known to be associated with different prognoses; for the latter, the prognosis is much poorer than for the former. The results of this study may at least partly explain this difference in prognosis.     

Biological monitoring of exposure to chlorpyrifos by high performance liquid chromatography

Sixteen patients with hypogonadotropic hypogonadism received gonadotropin replacement therapy. Two patients treated with HCG alone showed induction of spermatogenesis 2 and 12 months after the start of treatment. Three subjects receiving combination therapy showed sperm appearance 6-28 months after treatment. The patients showing sperm appearance, whose testicular volume was > or = 4 ml, showed a higher sperm count and impregnated their partners, although no relationship was found between pretreatment testicular volume and sperm appearance. The response to HCG test correlated with sperm appearance after gonadotropin therapy. Sperm appearance was not observed in any subject except for one who showed no response to luteinizing hormone-releasing hormone (LH-RH) test and none of the patients without response of FSH to LH-RH demonstrated any induction of spermatogenesis. In conclusion, the responses to LH-RH test and possibly to HCG test could predict the induction of spermatogenesis after gonadotropin replacement therapy, and a large testicular volume is associated with post-treatment fertility.     

Simultaneous determination of the binary mixture of nifedipine and mefruside using derivate spectroscopy, capillary gas-liquid chromatography and high performance liquid chromatography

Accurate, precise first-derivative ultraviolet spectrophotometric, gas-liquid and high performance liquid chromatographic methods for the determination of nifedipine and mefruside in tablet dosage form were described. The first-derivative method depends on measurements of the derivative amplitudes by peak-to-base and zero-crossing techniques, at 390 and 282 nm, for the determination of nifedipine and mefruside, respectively. Calibration graphs were linear (r = 0.9999) ranging from 8-40 and 20-60 micrograms ml-1 for nifedipine and mefruside, respectively, in presence of one another. The gas-liquid chromatographic method (GLC) was based on using cross-linked methylsilicone gum capillary column with flame ionization detector for the determination of nifedipine and mefruside in the binary mixture. The high performance liquid chromatographic (HPLC) method was based on using a reverse-phase column with a mobile phase of methanol-water (55-45, pH = 4.5) with UV detection at 260 nm. The three methods showed good linearity, precision and reproducibility. The proposed methods were successfully applied to the determination of both drugs in pharmaceutical tablets.     

Determination of free bile acids in pharmaceuticals by thin layer chromatography and high performance liquid chromatography

High-performance liquid chromatography with evaporative light scattering detection (HPLC-ELSD) and thin layer chromatography with flame ionization detection (TLC-FID) have been applied to the separation of five main free bile acids present in humans: cholic (CA), chenodeoxycholic (CDCA), deoxycholic (DCA), lithocholic (LCA) and ursodeoxycholic (UDCA) acid. HPLC separation was performed on Biospher Si 100 column using a mixture of n-heptane, isopropanol, ethylacetate, methanol and glacial acetic acid as a mobile phase. All the compounds were separated in less than 12 minutes by using a gradient elution mode. TLC-FID separation was performed on S-II Chromarods with a mixture of isooctane, ethylacetate and glacial acetic acid as a mobile phase. HPLC-ELSD method was applied to the determination of CDCA and UDCA in pharmaceuticals and their purity control when LCA, DCA and CA were considered as impurities.     

Separation from related compounds and assay of calcipotriol by high-performance liquid chromatography

In this paper, two methods are presented. One involves the separation of calcipotriol, a new synthetic analogue, from two related compounds, specifically cholecalciferol (Vitamin D3) and calcitriol (1,25-dihydroxyvitamin D3). The other involves the isolation and assay of calcipotriol from a topical ointment. The study was performed with reversed-phase high-performance liquid chromatography using an RP18 column and ultraviolet detection. Applying the method of Snyder, a mobile phase mixture containing methanol-acetonitrile-water (67:23:10, v/v) was found which achieved a total separation within 18 min. A mobile phase of methanol-water (80:20, v/v) attained a slower elution of calcipotriol. For isolation and assay of calcipotriol from an ointment (Daivonex), dissolution in chloroform gave the highest recovery (> 98%). The isolation and assay process can be performed within 2 h.     

Determination of nimesulide and hydroxynimesulide in human plasma by high performance liquid chromatography

Two specific methods for the simultaneous determination of nimesulide, a non steroidal anti-inflammatory drug, and its hydroxylated metabolite in human plasma are described. Adopting a high performance liquid chromatographic (HPLC) system with UV detection (230 nm), the compounds, extracted from plasma in acidic medium, were separated on ODS columns under gradient conditions, using a phosphate buffer solution and methanol as mobile phase. For each method column length, gradient rate and composition were appropriately selected. The limit of quantitation was 25 ng/mL for both compounds. The two methods were validated by intra day assays at three concentration levels and applied in kinetic studies in healthy volunteers, during which inter-day assays were carried out confirming their feasibility.     

Determination of provitamin A of green leafy vegetables by high performance liquid chromatography and open column chromatography

AIM OF STUDY: Intrathecal sufentanil has recently been used in labour as part of a combined spinal epidural technique. This study was conducted to compare its use in combination with bupivacaine for caesarean section with fentanyl added to bupivacaine and bupivacaine alone. METHODS: Sixty ASA I and II patients for non-emergency caesarean section under spinal anaesthesia were divided into three groups to receive 15 micrograms fentanyl added to 7.5 mg bupivacaine, 10 micrograms sufentanil added to 7.5 mg bupivacaine and 7.5 mg bupivacaine. Onset time of sensory blockade, side effects, surgical conditions, neonatal outcome and quality of the anaesthetic was assessed. On the first postoperative day, duration of effective analgesia, side effects and patient satisfaction were noted. RESULTS: The duration of effective analgesia of bupivacaine alone was prolonged with the addition of sufentanil and fentanyl by 358% and 256% respectively. No patient in the sufentanil and fentanyl groups required additional intra-operative analgesics compared with 17.6% of patients in the bupivacaine alone group. There was an increase in incidence of desaturation in the sufentanil group (45%) and fentanyl group (5.6%) compared with the bupivacaine only group (0%). The incidence of pruritus was 35% with sufentanil, 27.8% with fentanyl against 0% with bupivacaine alone. CONCLUSION: The addition of 10 micrograms of sufentanil and 15 micrograms of fentanyl to 7.5 mg of bupivacaine prolonged the duration of effective analgesia and improved intra-operative analgesia. However, the incidence of pruritus and episodes of desaturation were increased more with 10 micrograms sufentanil than with 15 micrograms fentanyl.     

Purification and initial characterization of the ATP:corrinoid adenosyltransferase encoded by the cobA gene of Salmonella typhimurium

The cobA gene of Salmonella typhimurium and its product were overexpressed to approximately 20% of the total cell protein. CobA was purified to 98% homogeneity; N-terminal sequence analysis (21 residues) of homogeneous protein confirmed the predicted amino acid sequence. ATP:corrinoid adenosyltransferase activity was demonstrated in vitro to be associated with CobA. This activity was optimal at pH 8 and 37 degrees C. A quantitative preference was determined for Mn(II) cations and ATP. The apparent Km of CobA for ATP was 2.8 microM, and that for cob(I)alamin was 5.2 microM. Vmax was measured at 0.43 nmol/min. Cobinamide served as the substrate for CobA to yield adenosylcobinamide. Activity was stable at 4 degrees C for several weeks but was lost rapidly at room temperature (50% overnight). Dithiothreitol was required to maintain the enzymatic activity of CobA.     

Purification and characterization of cysteine proteinase from a baculovirus gene

To analyze the degradation of product proteins at the late stage of virus infection in the baculovirus expression system, a cysteine proteinase was purified from hemolymph of Bombyx mori infected with wild-type B. mori nuclear polyhedorosis virus (BmNPV). The purified cysteine proteinase preparation had two protein bands (major 35-kDa active protein and 28-kDa inactive protein) on SDS-PAGE. Based on the N-terminal amino acid sequences of them, it was found that both proteins originated in the cysteine proteinase gene of BmNPV. The purified cysteine proteinase had an optimum pH at 4.0, and also had activities at neutral pHs. When recombinant luciferase was used as a natural substrate, it was degraded rapidly by the cysteine proteinase at the physiological pH of hemolymph. These results suggest that the cysteine proteinase from a BmNPV gene participates in the degradation of foreign protein expressed by the baculovirus system.     

A method for galactose-1-phosphate uridyltransferase assay and the separation of its isozymes by DEAE-cellulose column chromatography

The resistance to coronary blood flow in various parts of the myocardium was studied with the tracer microspheres technique before and immediately after an acute coronary occlusion and several weeks after a more slowly occurring coronary occlusion by Ameroid constrictor. All experiments were carried out in the isolated, metabolically supported, empty, beating dog heart at maximal coronary vasodilation induced with adenosine. Coronary resistance of the normal empty beating heart at maximal coronary vasodilation was 0.20 mm mm Hg/(ml/min) per 100 g of tissue (subepicardium) and 0.16 mm Hg/(ml/min) per 100 g of tissue (subendocardium). After acute coronary occlusion the perfusion of the subtended myocardium was maintained at a much lower level by way of collateral vessels, which showed a resistance to flow of 3.52 mm Hg/(ml/min) per 100 g. If coronary artery occlusion proceeded more slowly the collateral vessels became more functional and myocardial infarction was avoided. During collateral enlargement collateral resistance fell from 3.52 to 0.22 mm Hg/(ml/min) per 100 g within a period of 8 weeks after implantation of the constricting device. The degree of compensation by collaterals for the loss of the occluded native coronary artery was 33% of its former conductance.     

微柱高效液相色谱法测定环境样品中钴镍铜锌钒

合成了有机试剂2-(2-喹啉偶氮)-4-甲基-1,3-二羟基苯(QAMDHB),并以其为柱前衍生试剂,采用Waters XterraTMRP18(1.0 mm×50 mm,2.5μm)微色谱柱,φ=62%的甲醇(内含φ=0.5%的乙酸)为流动相,二极管阵列检测器,建立了高效液相色谱法测定环境样品中钴、镍、铜、锌和钒的新方法。根据信噪比(S/N=3)得各金属离子的检出限分别为:钴2μg/L,镍1.5μg/L,铜2μg/L,锌3μg/L,钒5μg/L,方法用于环境样品中痕量钴、镍、铜、锌、钒的测定,相对标准偏     

Retention behaviour of anti-emetic serotonin antagonists in reversed phase high performance liquid chromatography

Granisetron, Ondansetron and Tropisetron, three 5-HT3 antagonists showing anti-emetic activity, were analysed by HPLC on lipophilic stationary phases. The addition of an amine or quaternary ammonium salt to the eluents was a powerful tool in the analysis of these basic substances. The influence on chromatographic parameters of pH, ionic strength and various counter-ions in the aqueous phase as well as of different organic modifiers is discussed. Some of the proposed experimental conditions allow a more strictly partition-based separation mechanism and can produce chromatographic parameters suitable for structure-activity studies. These experimental conditions are also suitable for analysis of the considered compounds in pharmaceutical dosage forms or in biological fluids.