High-pressure Liquid chromatography-UV Research Articles

Residue Behavior of Clopyralid Herbicide in Soil and Sugar Beet Crop under Subtropical Field Conditions

Sugar beet is a major crop for the sugar industry. With growing awareness of unsystematic use of pesticides, health problem, and environmental issues, assessment of pesticide residues in soil and crops has become necessary. Studies of subtropical conditions on dissipation and residue analysis of clopyralid have not yet been reported. Therefore, dissipation kinetics and terminal residues of clopyralid for two cropping seasons in the soil and the sugar beet crop were studied under field conditions. An experiment was laid out in a randomized block design, and a herbicide was applied as a postemergent. Clopyralid was extracted from the matrix by basic water, subjected to solid phase extraction cleanup, and quantified by high-pressure liquid chromatography-UV. The method was validated, and recovery percentage of pesticide ranged from 81 to 88, 77 to 85, 78 to 86, and 89 to 94% in the soil, sugar beet roots, sugar beet leaves, and water, respectively. After application in the soil, clopyralid dissipated rapidly following monophasic first-order kinetics, with a half-life of 13.39 days. Limits of detection and quantitation were 0.007 and 0.02 μg g-1, respectively. Clopyralid does not persist long in soil, and residues were below the European Union's maximum residue levels (0.5 mg kg-1) in the roots and leaves of sugar beet. Residues were also not detected in the groundwater. It can be concluded that clopyralid could be considered a safe herbicide from the environmental aspect due to its nonpersistence and that it would not have an adverse effect on human or animal health.

Evaluation of the association of SNP in carboxylesterase enzyme (CES1) with pharmacokinetic and adverse effects of capecitabine in breast and colorectal cancer patients

Capecitabine is an oral prodrug and converted to 5-fluorouracil using three-step enzymatic pathways which include carboxylesterase (CES). Interindividual differences in the activities of drug-metabolizing enzymes may affect efficacy and toxicity. The aim of this study is to evaluate the association of Single nucleotide polymorphisms (SNP) in CES1 with the pharmacokinetic and adverse effects of capecitabine. Plasma samples were obtained from 7 breast and colorectal cancer patients who were treated with capecitabine-based chemotherapy (1000-1250 mg/m2 ) at 0.5, 1, 2, 3 and 4 hours following drug administration on their first day of the first cycle. The plasma concentrations of the capecitabine were determined by using a high-pressure liquid chromatography-UV detector. SNP (rs8192950) was genotyped using the reverse transcription-polymerase chain reaction. Patients were found to have heterozygote (57%), wild (29%), and mutant (14%) distributions of genotypes (p=0.909). The mean plasma area under the curve (AUC0-4h) was 4.60±2.25 µg.h/mL, and maximum plasma concentration (Cmax) was 3.19±2.5 µg/mL. There were no statistically significant differences between genotypes and AUC values (p=0.2236) and the most frequently observed side effects were diarrhea (p=0.1028), asthenia (p=0.6456), anemia (p=0.6456), emesis (p=0.3499). This is the first study evaluating an association of genetic variation in CES1 (rs8192950) with pharmacokinetic and adverse effects of capecitabine. Therefore, additional study in larger groups of patients is required to support our study. 

Production of chlorzoxazone glucuronides via cytochrome P4502E1 dependent and independent pathways in human hepatocytes.

CYP2E1 activity is measured in vitro and in vivo via hydroxylation of the Chlorzoxazone (CHZ) producing the 6-hydroxychlorzoxazone (OH-CHZ) further metabolized as a glucuronide excreted in urine. Thus, the quantification of the OH-CHZ following enzymatic hydrolysis of CHZ-derived glucuronide appears to be a reliable assay to measure the CYP2E1 activity without direct detection of this glucuronide. However, OH-CHZ hydrolyzed from urinary glucuronide accounts for less than 80% of the CHZ administrated dose in humans leading to postulate the production of other unidentified metabolites. Moreover, the Uridine 5'-diphospho-glucuronosyltransferase (UGT) involved in the hepatic glucuronidation of OH-CHZ has not yet been identified. In this study, we used recombinant HepG2 cells expressing CYP2E1, metabolically competent HepaRG cells, primary hepatocytes and precision-cut human liver slices to identify metabolites of CHZ (300μM) by high pressure liquid chromatography-UV and liquid-chromatography-mass spectrometry analyses. Herein, we report the detection of the CHZ-O-glucuronide (CHZ-O-Glc) derived from OH-CHZ in culture media but also in mouse and human urine and we identified a novel CHZ metabolite, the CHZ-N-glucuronide (CHZ-N-Glc), which is resistant to enzymatic hydrolysis and produced independently of CHZ hydroxylation by CYP2E1. Moreover, we demonstrate that UGT1A1, 1A6 and 1A9 proteins catalyze the synthesis of CHZ-O-Glc while CHZ-N-Glc is produced by UGT1A9 specifically. Together, we demonstrated that hydrolysis of CHZ-O-Glc is required to reliably quantify CYP2E1 activity because of the rapid transformation of OH-CHZ into CHZ-O-Glc and identified the CHZ-N-Glc produced independently of the CYP2E1 activity. Our results also raise the questions of the contribution of CHZ-N-Glc in the overall CHZ metabolism and of the quantification of CHZ glucuronides in vitro and in vivo for measuring UGT1A activities.

Application of HPLC (High Pressure Liquid Chromatography) for Analysis of Lard in the Meatball Product Combined with PCA (Principal Component Analysis)

Meatball is favorite food for Indonesian society. Nowadays, many issues about the food ingredient on meatball were mixed with Lard. Therefore, this study was focused on applications of HPLC UV for the lard detection in meatball. The results showed that the beef and pork meatball can be distinguished using HPLC which was combined Principal Component Analysis (PCA). However, the application of HPLC has a problem for lard detection in meatball because can’t to control the hydrolysis of triglyceride (TGA). Hydrolysis of triglycerides can also be detected using HPLC UV and giving the interference for halal authentication.

Influence of Thawing Methods and Storage Temperatures on Bacterial Diversity, Growth Kinetics, and Biogenic Amine Development in Atlantic Mackerel

Limited knowledge is currently available on the influence of fish thawing and subsequent storage conditions on bacterial growth kinetics, succession, and diversity alongside the production of biogenic amines. This study aimed to address these factors during the thawing and subsequent storage of mackerel. Thawing was either done fast in 18°C water for 2 h or slowly at 30°C overnight. Subsequent storage was at 30°C (ambient) for 36 h and 2 to 5°C (refrigerated) for 12 days. The cultivation methods used were total viable counts, hydrogen sulfide-producing bacteria, and Pseudomonas . Maximum growth rate, population density, and lag time were fitted on the counts using the Baranyi model. The bacterial diversity and succession were based on sequencing of 16S rRNA amplicons, and biogenic amines were quantified on high-pressure liquid chromatography-UV. The results show that lag time of hydrogen sulfide-producing bacteria was significantly affected by both thawing methods, and further, the interaction between thawing and storage significantly affected the maximum growth rate of these bacteria. However, the maximum growth rate of Pseudomonas was higher during refrigerated storage compared with storage at ambient temperature. Total viable counts showed longer lag time and reduced growth rate under refrigerated storage. Higher bacterial diversity was correlated to slow thawing and storage at ambient temperature compared with slow thawing and refrigerated storage. Overall, Acinetobacter and Psychrobacter genera were the dominant bacterial populations. The amine levels were low and could not be differentiated along the thawing and storage approaches, despite a clear increase in bacterial load, succession, and diversity. This corresponded well with the low abundance of biogenic amine-producing bacteria, with the exception of the genus Proteus , which was 8.6% in fast-thawed mackerel during storage at ambient temperature. This suggests that the decarboxylation potential is dependent on both microbial load and microbial community structure.

Treatment of human and livestock helminth infections in a mobile pastoralist setting at Lake Chad: Attitudes to health and analysis of active pharmaceutical ingredients of locally available anthelminthic drugs

Mobile pastoralists face challenges in accessing quality health care and medication for managing human and animal diseases. We determined livestock disease priorities, health seeking behaviour of people bearing helminthiases and − placing particular emphasis on trematode infections − treatment strategies and outcome satisfaction among mobile pastoralists of four ethnic groups in the Lake Chad area using focus group discussions. People suffering from schistosomiasis were interviewed about symptoms, health seeking behaviour and their satisfaction with respect to the provided treatment. Anthelminthic drugs for human and veterinary use obtained from various health care structures were analysed for active pharmaceutical ingredients (API) and quantity, using high pressure liquid chromatography-UV and liquid chromatography combined with tandem mass spectrometry. Most people suffering from schistosomiasis sought treatment at health care centres. Yet, they also consulted informal providers without medical training. Regarding animal health, self-mediated therapy was common to manage suspected livestock fascioliasis. Self-reported treatment satisfaction for human schistosomiasis and trematodiasis treatment outcome in livestock were low. Mobile pastoralists perceived the purchased drugs to be of low quality. Among 33 products locally sold as anthelminthic drugs for human or veterinary use, 27 contained albendazole or mebendazole, varying between 91% and 159% of the labelled amount. Six products were sold loosely with incomplete information and their API could not be identified. No counterfeit anthelminthic drugs were detected. None of the samples contained praziquantel or triclabendazole, the drugs of choice against schistosomiasis and fascioliasis, respectively. The perceived unsatisfactory treatment outcomes in humans and animals infected with trematodes are most likely due to empiric diagnosis and the resulting use of inadequate therapy for human schistosomiasis and the lack of efficacious drugs against livestock fascioliasis.

Development of Budesonide Nanocluster Dry Powder Aerosols: Formulation and Stability

The physical and chemical stability of dry powder aerosol formulations is an essential component in the development of an inhaled therapeutic. The pharmaceutical processing methods and storage conditions are primary determinants of the stability of a dry powder inhaler (DPI) formulation. Wet milling was used to produce budesonide NanoClusters (NCs), which are agglomerates of drug nanoparticles (≈ 300 nm) with a mean aerodynamic diameter between 1 and 3 µm, capable of deep lung penetration. In this study, the reproducibility of NC processing and performance was established. The physical stability of a selected budesonide NC formulation was investigated using industry standard dose content uniformity and cascade impaction techniques. The chemical stability of the lead formulation was also determined as a function of processing parameters and storage conditions. This study confirms the reproducibility and robust stability of NC powders as a novel means to turn drug particles into high-performance aerosols.

Pilot Study of Oral Silibinin, a Putative Chemopreventive Agent, in Colorectal Cancer Patients: Silibinin Levels in Plasma, Colorectum, and Liver and Their Pharmacodynamic Consequences

Silibinin, a flavonolignan from milk thistle, has intestinal cancer chemopreventive efficacy in rodents. It is a strong antioxidant and modulates the insulin-like growth factor (IGF) system by increasing circulating levels of IGF-binding protein 3 (IGFBP-3) and decreasing levels of IGF-I. Here, the hypothesis was tested that administration of oral silibinin generates agent levels in human blood and colorectal and hepatic tissues consistent with pharmacologic activity. Patients with confirmed colorectal adenocarcinoma received silibinin formulated with phosphatidylcholine (silipide) at dosages of 360, 720, or 1,440 mg silibinin daily for 7 days. Blood and biopsy samples of normal and malignant colorectum or liver were obtained before dosing, and blood and colorectal or hepatic tissues were collected at resection surgery after the final silipide dose. Levels of silibinin were quantified by high-pressure liquid chromatography-UV, and plasma metabolites were identified by liquid chromatography-mass spectrometry. Blood levels of IGFBP-3, IGF-I, and the oxidative DNA damage pyrimidopurinone adduct of deoxyguanosine (M1dG) were determined. Repeated administration of silipide was safe and achieved levels of silibinin of 0.3 to 4 micromol/L in the plasma, 0.3 to 2.5 nmol/g tissue in the liver, and 20 to 141 nmol/g tissue in colorectal tissue. Silibinin monoglucuronide, silibinin diglucuronide, silibinin monosulfate, and silibinin glucuronide sulfate were identified in the plasma. Intervention with silipide did not affect circulating levels of IGFBP-3, IGF-I, or M1dG. The high silibinin levels achieved in the human colorectal mucosa after consumption of safe silibinin doses support its further exploration as a potential human colorectal cancer chemopreventive agent.

Systemic availability of prophylactic cefuroxime in patients submitted to coronary artery bypass grafting with cardiopulmonary bypass

Cardiopulmonary bypass and hypothermia (HCPB) is a procedure commonly used during heart surgery, representing a risk factor for the patient by promoting extensive haemodilution and profound physiological changes. Cefuroxime is used for the prevention of infection following heart surgery, and several dose schemes have been suggested for prophylaxis with cefuroxime. The objective of the present study was to assess, in a comparative manner, the systemic availability of cefuroxime administered intravascularly as a bolus dose of 1.5 g to 17 patients having heart surgery with or without HCPB. Plasma cefuroxime concentrations were determined by high-pressure liquid chromatography-UV, and the following values, expressed as medians, were obtained for the study group compared with controls: 69.1 vs. 62.7 mg/L (1st h), 35.8 vs. 26.0mg/L (3rd h), 14.6 vs. 8.7 mg/L (6th h, P<0.05), 6.1 vs. 3.0mg/L (9th h, P<0.05) and 2.6 vs. 1.0mg/L (12th h, P<0.05). Despite the differences recorded during the study period as a consequence of HCPB, low antibiotic concentrations were found as early as 6h post dose for both groups investigated. Thus, the low systemic availability of cefuroxime after the administration of a 1.5-g dose may not protect against postoperative infections. The data obtained permit us to recommend a change in the dose scheme in order to maintain adequate plasma levels of cefuroxime.

Temporal variation of tannins (galloylglucoses), flavonols and anthocyanins in leaves of Eucalyptus nitens seedlings: implications for light attenuation and antioxidant activities

Leaves of Eucalyptus nitens (Deane &amp; Maiden) Maidenwere sampled during establishment of seedlings that were deprived of nutrientsor fertilised at planting. Extracts were analysed for polyphenolic compoundsusing high pressure liquid chromatography (HPLC) in conjunction with UV diodearray and electrospray ionisation mass spectrometry (ESI-MS) with negative iondetection as well as direct MS injection. The presence of quercetin, rutin,sideroxylonal A and B andpenta-galloylglucose were confirmed by comparison with standards. Othergalloylglucoses were identified by first principles. Galloylglucoses comprisedabout 75% of polyphenolics. Quercetin and various quercetin glycosidescomprised ca 85% of flavonols. The anthocyanins,cyanidin-3-glycoside and cyanidin-3,5-diglycoside were identified from directinjection MS data. HPLC–ESI-MS was found to be a superior analyticaltechnique compared to conventional HPLC–UV diode array detection due toco-elution of peaks under UV. Temporal variation of anthocyanins may berelated to the severity of cold-induced photoinhibition. Changes ingalloylglucoses and flavonols were similar with time and related to levels offoliar nitrogen. The suitability of HPLC–ESI-MS for the study of foliarpolyphenolics, the phenolic constitution of E. nitensfoliage, and temporal variation of the phenolic constituents during a seedlingestablishment period of 29 weeks are demonstrated.

A Preliminary Survey of Antibiotic Residues and Viable Bacteria in Milk from Three Caribbean Basin Countries

There is widespread concern about the presence of antimicrobial drugs in milk. The presence of drug residues in milk may have public health implications. Milk samples (n = 25 to 65/country) were collected from bulk tanks and commercial vendors in Barbados, Costa Rica, and Jamaica between February 1996 and August 1997. Bulk tank samples were collected from high milk-producing regions of Jamaica and Costa Rica and from 26 dairy farms in Barbados. Milk pH, bacterial growth (total CFU/ml and the presence of Streptococcus agalactiae and Staphylococcus aureus), and the presence of antimicrobials were determined. Milk samples were tested by a microbial inhibition test (Delvotest-P, Gist-Brocades Food Ingredients, Inc.) to screen for antimicrobial drugs. All positives were retested for the presence of β-lactam antibiotics after incubating with penicillinase and some positives were identified by high-pressure liquid chromatography–UV. Mean pH values ranged from 6.5 to 6.7. S. aureus was identified in bulk tank samples from Costa Rica (52%), Barbados (44%), and Jamaica (46%). S. agalactiae was identified in bulk tank samples from Costa Rica (28%), Barbados (8 and 16%), and Jamaica (18%). Antimicrobial residues were detected in some bulk tank samples from Barbados (8%) and Jamaica (10%) but not in samples from Costa Rica. All positives in milk from Jamaica and Barbados were determined to be β-lactams. No residues were detected in pasteurized milk samples from Barbados or ultrahigh-temperature milk from Jamaica. The presence of β-lactam residues in some of these samples suggests the appropriateness of testing milk prior to processing for consumption.

Excretion of cefprozil into human breast milk.

The excretion of cefprozil into breast milk in nine healthy, lactating female subjects was investigated. Each subject received a single 1,000-mg oral dose of cefprozil consisting of cis and trans isomers in an approximately 90:10 ratio. Serial blood, urine, and breast milk samples were collected and analyzed for the concentrations of the cis and trans isomers by a specific high-pressure liquid chromatography-UV assay. The mean pharmacokinetic parameters for both isomers were essentially the same. The mean peak concentrations in plasma for the cis isomer were 14.8 micrograms/ml, and the area under the concentration curve was 54.8 micrograms.h/ml. The mean values of elimination half-life, renal clearance, and urinary excretion for the cis isomer were 1.69 h, 164 ml/min, and 60%, respectively. The mean concentrations in milk of the cis isomer over a 24-h period ranged from 0.25 to 3.36 micrograms/ml, with the maximum concentration appearing at 6 h after dosing. The average maximum concentration in milk of the trans isomer was less than 0.26 micrograms/ml. The concentrations of the trans isomer in plasma and in breast milk were about 1/10 of those for the cis isomer. Less than 0.3% of the dose was excreted in breast milk for both isomers of cefprozil. Even if one assumes that the concentration of cefprozil in milk remains constant at 3.36 micrograms/ml (the highest concentration of cefprozil observed in breast milk), an infant ingesting an average of 800 ml of milk per day will be exposed to a maximum amount of about 3 mg of cefprozil per day. This value represents about 0.3% of the maternal dose. Low excretion of cefprozil in breast milk and the excellent safety profile of cefprozil suggest that this cephalosporin may be administered to nursing mothers when indicated.

Formation of 8-methylguanine as a result of DNA alkylation by methyl radicals generated during horseradish peroxidase-catalyzed oxidation of methylhydrazine.

Methylhydrazine oxidation promoted by horseradish peroxidase-H2O2 or ferricyanide led to the generation of high yields of methyl radicals and to the formation of 7-methylguanine and 8-methylguanine upon interaction with calf thymus DNA. Methyl radicals were identified by spin-trapping experiments with alpha-(4-pyridyl-1-oxide)-N-tert-butyl nitrone and tert-nitrosobutane. The methylated guanine products were identified in the neutral hydrolysates of treated DNA by high pressure liquid chromatography (HPLC) analysis and spiking with authentic samples. The structure of 8-methylguanine, a product not previously reported in enzymatic systems, was confirmed by HPLC chromatography, UV absorbance, and mass spectrometry. The formation of 8-methylguanine suggests a possible role for carbon-centered radicals as DNA-alkylating agents.

Pharmacokinetics of the novel cephalosporin cefepime (BMY-28142) in rats and monkeys.

The disposition of the novel cephalosporin cefepime (BMY-28142) was characterized for intravenous administration of single doses to rats and cynomolgus monkeys, the species used most extensively for safety evaluation of the compound. Serial blood samples were collected from individual animals, and plasma was analyzed for intact cefepime by a high-pressure liquid chromatography-UV method. Assay results were evaluated by compartmental and noncompartmental methods to characterize pharmacokinetics for each species and dosage regimen. For intravenous (i.v.) bolus administration of 28 to 386 mg/kg (body weight) to rats, total body clearance (CL; 11.0 ml/min per kg) was essentially invariant with the dose; however, the terminal half-life (t1/2) and the steady-state distribution volume (Vss) increased with increasing dose level. After administration of 87 to 1,502 mg/kg by i.v. infusion, CL (12.5 ml/min per kg) was again similar for all dose groups. Mean t1/2 values (1.3 to 4.6 h) appeared unusually long for a cephalosporin in rats, and inordinately variable. No consistent differences among dose group mean Vss values were found. The maximal concentration of drug in plasma at the end of infusion was not a linear function of dose. For the cynomolgus monkey, kinetic parameters for 5-min i.v. infusions were linearly related to dose over the range of 10 to 600 mg/kg. Mean parameter values were t1/2 = 1.7 h, CL = 1.6 ml/min per kg, and Vss = 0.21 liters/kg. The pharmacokinetic results indicate substantive differences between the two species with respect to their response to toxicologic doses of cefepime.

Glomerular arachidonate lipoxygenation in rat nephrotoxic serum nephritis.

Arachidonate lipoxygenation to monohydroxylated eicosatetraenoic acids (HETE) was studied in rat nephrotoxic serum nephritis (NSN). A single infusion of nephrotoxic serum enhanced conversion of [3H]arachidonic acid ([3H]C20:4) to [3H]12-HETE in glomeruli isolated from nephritic rats compared with controls. The percent conversion of [3H]arachidonic acid was 1.95 +/- 0.2% in control glomeruli and 14.2 +/- 2% in nephritic glomeruli 2 d after induction of disease. No significant changes in the conversion of [3H]C20:4 to [3H]5-, 8-, and 9-HETE were noted. Extraction of glomerular HETE by alkaline hydrolysis, to evaluate possible reacylation of HETE after their production, confirmed the presence of 12-HETE and did not provide evidence of 5-HETE synthesis. Increased glomerular 12-HETE synthesis in nephritic rats was also demonstrated by high pressure liquid chromatography-UV detection and by 12-HETE radioimmunoassay. The enhanced glomerular 12-HETE synthesis commenced as early as 3-5 h after administration of nephrotoxic serum and peaked at day 2 with 10-fold enhancement of 12-HETE production. Increments of glomerular 12-HETE persisted on day 7 and returned toward control levels by day 14. Platelet depletion, induced by antiplatelet antisera, did not decrease glomerular 12-HETE synthesis in NSN, thereby eliminating platelets as the cellular origin of 12-HETE. Glomerular epithelial and mesangial cells are the most likely sources of enhanced 12-lipoxygenase activity. The enhanced arachidonate 12-lipoxygenation in glomerular immune injury could have important proinflammatory effects in the evolution of glomerulonephritis since 12-HETE has important effects on leukocyte function.