Piromidic Acid Research Articles

Simultaneous determination of quinolones in livestock and marine products by high performance liquid chromatography

A high performance liquid chromatographic (HPLC) method was developed for the simultaneous determination of nalidixic acid, flumequine, oxolinic acid, piromidic acid, danofloxacin, enrofloxacin and sarafloxacin in chicken, pork and fish. These seven quinolone antibacterials were extracted with 0.3% metaphosphoric acid: acetonitrile (1:10, v/v), followed by a Bond Elut C18 cartridge clean up procedure. The HPLC separation was achieved on a Cosmosil 5C18-AR-II column (5 μm, 4.6 mm i.d. × 250 mm) with acetonitrile: 0.05M NaH2PO4 (pH 2.5)(35:65, v/v) containing 3.5 mM sodium dodecyl sulfate as a mobile phase, and detection was performed with photodiode array and fluorescence detector (by using wavelength programming). Good linearity was observed from the calibration plot at concentrations from 0.05 to 10.0 mg/mL (0.005-2.0 μg/mL for danofloxacin). Recovery studies of the analytes were performed at 0.01, 0.1, 0.4 and 2.0 ppm (0.001, 0.01, 0.04 and 0.20 ppm for danofloxacin) spiked levels. Average recoveries of low concentration ranged from 74.3 to 85.5% and those of the rest ranged from 80.1 to 99.9% with coefficients of variation less than 5.8%. The detection limits of quinolones were 0.01-0.04 ppm with photodiode array detection and 0.0006-0.05 ppm with fluorescence detection. The coefficients of variation of intra-day and inter-day assays were lower than 3.29% and 5.23%, respectively. These results indicated that the developed method had an acceptable precision. Using this method to detect quinolones in sixty three samples purchased from various markets in Taipei, we found that nine wu ku chicken muscles contained enrofloxacin residues ranged from 0.08 to 4.04 ppm, four wu ku chicken liver muscles contained enrofloxacin residues ranged from 0.01 to 0.27 ppm, and oxolinic acid residues in three sweet fish samples ranged from 0.13 to 0.35 ppm. The results indicated that 25.4% of samples violated the regulation set by the Department of Health. The enrofloxacin residue in wu ku chicken muscle is very stable even after a long period of refrigeration at 4°C.

Analysis of aristolochic acid in asarum (Xixin) and its preparations by liquid chromatography/tandem mass spectrometry

A method for analysis of aristolochic acids (AAs: AA-I, AA-II) in Asarum (Xixin, named in Chinese) and its preparations by liquid chromatography coupled with tandem mass spectrometry has been developed. The chromatographic analysis was carried out using a C18 column (2.1x150 mm) and a mixture of 0.1% formic acid (containing 0.1% ammonium acetate) and acetonitrile (65:35, v/v) as the mobile phase at a flow rate of 0.3 mL/min. Piromidic acid was used as an internal standard. The mass detection was measured as the positive ion electrospray ionization mode. The precursor and daughter ions for quantitation were m/z 359 and 298 for AA-I, m/z 329 and 268 for AA-II. The instrument detection limit of AA-I and AA-II was 2 and 2.8 ng/mL, respectively. The result showed that Asarum contained AA-I while AA-II was not detected. AA-I distribution in different parts of Asarum was examined in detail and AA-I content in both methanol and water extracts was also compared in this study. In our twenty samples of Asarum, the average amounts of AA-I in 75% methanol extracts of roots, rhizomes, petioles and leaves were 1.23 ± 0.64, 1.72 ± 1.38, 7.81 ± 4.04 and 27.13 ± 25.31 ppm, respectively. The average amounts of AA-I in water extracts of roots, rhizomes, petioles and leaves were 0.33 ± 0.32, 0.45 ± 0.50, 3.98 ± 3.09 and 6.32 ± 9.75 ppm, respectively. The results indicated that the amounts of AA-I in roots were less than those in petioles and leaves, and the amounts of AA-I in water extracts were about 25 to 50 percent of those in 75% methanol extracts. For Asarum preparations, the amounts of AA-I in each six of Xi-xin-extract- powder and Ma-Huang-Fu-Zi-Xi-Xin-Tang ranged between 0.35-2.07 ppm and 0.21-1.00 ppm, respectively. Results of this study could be provided for the regulatory authority as references.

Simultaneous determination of 18 quinolone residues in marine and livestock products by liquid chromatography/tandem mass spectrometry

A simple and efficient LC/ESI/MS/MS method was developed to determine 18 (fluoro)quinolone (QNs) residues in milk, chicken, pork, fish and shrimp. This method is capable of screening and confirming the presence of 12 amphoteric QNs (marbofloxacin, norfloxacin, enrofloxacin, ciprofloxacin, desethylene ciprofloxacin, lomefloxacin, danofloxacin, sarfloxacin, difloxacin, ofloxacin, orbifloxacin and enoxacin) and 6 acidic QNs (oxolinic acid, nalidixic acid, flumequine, cinoxacin, piromidic acid and pipemidic acid). The drugs were extracted from matrix with acetonitrile containing 1% formic acid, diluted in 10% acetonitrile and defatted by extraction with hexane. The LC separation was conducted on an XDB C-8 (150 mm x 4.6 mm, 5 μm) column with gradient elution of 20 mM ammonium formate in 0.1% formic acid-acetonitrile as the mobile phase. Mass spectrum acquisition was completed in the positive ion mode by applying multiple reaction mode (MRM). The decision limit (CCα) and detection capability (CCβ), stated in the Decision No. 2002/657/EC and the ISO standard No.11843, have been calculated in the case of nonauthorized substance. The values of CCα ranged from 0.18 to 0.68 ng/g and CCβ ranged from 0.24 to 0.96 ng/g under specified conditions.

Assessing physical properties of amphoteric fluoroquinolones using phosphorescence spectroscopy.

The self-association of fluoroquinolones (FQ) in water would play a relevant role in their translocations across lipid membranes. Triplet excited states of these drugs have been shown as reporters of FQ self-association using laser flash photolysis technique. A study using low-temperature phosphorescence technique was performed with quinolone derivatives such as enoxacin (ENX), norfloxacin (NFX), pefloxacin (PFX), ciprofloxacin (CPX, ofloxacin (OFX), nalidixic acid (NLA), pipemidic acid (PPA) and piromidic acid (PRA) to explore emission changes associated with self-associations and to shed some light on the triplet excited state energy (ET) discrepancies described in the literature for most of these drugs. The emissions obtained at 77 K in buffered aqueous medium revealed that the amphoteric nature of the quinolones CPX, NFX, PFX, ENX, OFX and PPA must generate their self-associations because a redshift of their phosphorescence maxima is produced by FQ concentrations increases. Hence, this effect was not observed for NLA and PRA or when all quinolones were analysed using ethanol or ethylene glycol aqueous mixtures as glassed solvents. Interestingly, the presence of these organic mixtures produced a blue-shift in the phosphorescence emission maximum of each FQ. Additionally, laser flash photolysis experiments with PRA and the amphoteric quinolone PPA, compounds with the same skeleton but different peripheral substituent, confirm the expected correlations between the amphoteric nature of compounds and their self-associations in aqueous media because the excimer generation was only detected for PPA. Now, the discrepancies described in the literature for the ET of FQs can be understood considering that changes of medium polarity or proticity as well as the temperature can considerably modify their ET values. Thereby, low-temperature phosphorescence technique, is an effective way to detect molecular self-associations and surrounding changes in quinolones that opens the possibility to evaluate these effects in other drug families.

Determination of quinolone residues in raw cow milk. Application of polar stir-bars and ultra-high performance liquid chromatography–tandem mass spectrometry

ABSTRACTSeventeen quinolone antibiotics were determined in cows’ milk. A method of high sensitivity, selectivity and accuracy was developed. Accuracy (trueness and precision), linearity, sensitivity, selectivity, decision limit and detection capability were established following the recommendations of the Commission Decision 2002/657/EC and the Food and Drug Administration (FDA) guideline. The use of polar stir-bar sorptive extraction (SBSE) prior to UHPLC-MS/MS analysis is proposed. The variables that affect SBSE were optimised using multivariate optimisation strategies. The ionic strength, the extraction time and the sample volume were studied. pH and stir-bar coating (polydimethylsiloxane, PDMS, and polyethyleneglycol modified silicone, PEG) were studied. PEG showed the best extraction yield at pH 6. For validation, a matrix-matched calibration and a recovery assay were carried out. Limits of quantification from 0.5 μg kg−1 for nalidixic acid, flumequine and piromidic acid, to 4.0 μg kg−1 for sarafloxacin were calculated. The precision (%, RSD) was lower than 15% for all antibiotics. Recoveries in fortified samples were between 88 and 114%.

Analytical methods for the determination of some selected 4-quinolone antibacterials

AbstractA comprehensive review with 337 references for the analysis of some selected 4-quinolone drugs belonging to the first and second generations since 2006 up till now is presented. This group includes nalidixic acid, oxolinic acid, piromidic acid, pipemidic acid and rosoxacin from the first generation and enoxacin, fleroxacin, nadifloxacin, pefloxacin and rufloxacin from the second generation. The review covers most of the methods described for the analysis of these drugs, either

Gold nanoparticle-based paper sensor for ultrasensitive and multiple detection of 32 (fluoro)quinolones by one monoclonal antibody

For rapid and simultaneous detection of (fluoro)quinolones, a broadly specific monoclonal antibody (mAb) that recognizes 32 (fluoro)quinolone antibiotics was prepared using a mixture of a norfloxacin derivative and a sarfloxacin derivative as the hapten. An immunochromatographic strip based on gold nanoparticles (AuNPs) was then assembled with goat anti-mouse antibody and antigen (sarfloxacin coupled to ovalbumin), used to form the C line and T line, respectively. This antigen competes with the (fluoro)quinolones in a sample incubated with mAbs labeled with AuNPs. The strip can detect 32 (fluoro)quinolones including oxolinic acid, nalidixic acid, miloxacin, pipemidic acid, piromidic acid, rosoxacin, cinoxacin, norfloxacin, pefloxacin, lomfloxacin, enofloxacin, fleroxacin, ciprofloxacin, enrofloxacin, dafloxacin, orbifloxacin, sparfloxacin, gemifloxacin, besifloxacin, balofloxacin, gatifloxacin, moxifloxacin, nadifloxacin, ofloxacin, marbofloxacin, flumequine, pazufloxacin, prulifloxacin, sarafloxacin, difloxacin, trovafloxacin, and tosufloxacin in milk within 10 min with the naked eye. The cut-off values of the strip range from 1 to 100 ng/mL and the limits of detection are 0.1–10 ng/mL. The strip does not cross-react with antibiotics including tetracycline, sulfamethazine, ampicillin, erythromycin, aflatoxin B1, or gentamicin. In short, this immunochromatographic strip is a very useful tool for the primary screening of (fluoro)quinolones in milk.

Removal of quinolone antibiotics from wastewaters by sorption and biological degradation in laboratory-scale membrane bioreactors

Laboratory-scale batch experiments were developed to investigate the main removal routes for 6 commonly found quinolones (ciprofloxacin, moxifloxacin, norfloxacin, ofloxacin, pipemidic acid, and piromidic acid), in wastewaters from a wastewater treatment plant, at μgL−1 levels in an aerobic sludge system from a membrane bioreactor (MBR) pilot plant. It was demonstrated that sorption and biotransformation were the main removal routes for the target antibiotics over other possible pathways, as volatilization or hydrolysis, under the experimental conditions. Mass balances indicated that sorption on sludge played a dominant role in the elimination of antibiotics from waters. The sorption coefficient Kd depended strongly on temperature and on the quinolone type and were higher at lower temperatures and for piperazinylic quinolones. Kd values were between 516 and 3746Lkg−1 in the temperature range of 9–38°C. Higher mixed liquor suspended solids (MLSS) increased quinolone removal efficiency mainly by sorption. Quinolone biodegradation constituted a secondary pathway, and could be described by first-order kinetics with degradation-rate constants ranging from 8.0×10−4h−1 to 1.4×10−2h−1 within the same temperature range and MLSS from 7000 to 15,000mgL−1. Biodegradation depended on the MLSS and temperature, but also on the initial chemical oxygen demand (COD). Higher biodegradation rates were observed at higher MLSS and temperature, as well as at low initial COD. Ciprofloxacin and moxifloxacin registered the highest biodegradation percentages (52.8% and 47.2%, respectively, at 38°C and 15,000mgL−1 MLSS), which is evidence that, despite the known persistence of this group of antibiotics and removal from waters mainly by sorption, it was possible to improve their removal by biodegradation, with an appropriate selection of conditions and control of process variables, as a preliminary step towards the elimination of these antibiotics from the environment. Further research is needed on the possibilities of removing sorbed antibiotics from sludge.

Laser diode thermal desorption mass spectrometry for the analysis of quinolone antibiotic residues in aquacultured seafood

Veterinary drug residue analysis of meat and seafood products is an important part of national regulatory agency food safety programs to ensure that consumers are not exposed to potentially dangerous substances. Complex tissue matrices often require lengthy extraction and analysis procedures to identify improper animal drug treatment. Direct and rapid analysis mass spectrometry techniques have the potential to increase regulatory sample analysis speed by eliminating liquid chromatographic separation. Flumequine, oxolinic acid, and nalidixic acid were extracted from catfish, shrimp, and salmon using acidified acetonitrile. Extracts were concentrated, dried onto metal sample wells, then rapidly desorbed (6 s) with an infrared diode laser for analysis by laser diode thermal desorption atmospheric pressure chemical ionization with tandem mass spectrometry (LDTD-MS/MS). Analysis was conducted in selected reaction monitoring mode using piromidic acid as internal standard. Six-point calibration curves for each compound in extracted matrix were linear with r(2) correlation greater than 0.99. The method was validated by analyzing 23 negative samples and 116 fortified samples at concentrations of 10, 20, 50, 100, and 600 ng/g. Average recoveries of fortified samples were greater than 77% with method detection levels ranging from 2 to 7 /g. Three product ion transitions were acquired per analyte to identify each residue. A rapid method for quinolone analysis in fish muscle was developed using LDTD-MS/MS. The total analysis time was less than 30 s per sample; quinolone residues were detected below 10 ng/g and in most cases residue identity was confirmed. This represents the first application of LDTD to tissue extract analysis. Published 2012. This article is a US Government work and is in the public domain in the USA.

Piromidic acid dry syrupによる小児細菌性下痢症の治療

Piromidic acid dry syrupによる小児細菌性下痢症の治療

조피볼락, Sebastes schlegeli에 경구투여된 nalidixic acid 및 piromidic acid의 약물동태에 미치는 수온의 영향

Nalidixic acid(NA), piromidic acid(PA)를 사육수온 (<TEX>$13{\pm}1.5^{\circ}C$</TEX>, <TEX>$23{\pm}1.5^{\circ}C$</TEX>)에 따라서 조피볼락(평균체중 <TEX>$500{\pm}30\;g$</TEX>)에 60 mg/kg의 농도로 1회 경구투여한 다음, 시간경과에 따라 혈청 내 잔류농도를 분석하였다. <TEX>$23{\pm}1.5^{\circ}C$</TEX>의 경우, 투여 후 NA는 24시간째(<TEX>$5.87\;{\mu}g/ml$</TEX>), PA도 24시간째(<TEX>$0.43\;{\mu}g/ml$</TEX>)에 각각 최대혈중농도에 도달하였다. <TEX>$13{\pm}1.5^{\circ}C$</TEX>의 경우, 투여 후 NA는 10시간 째(<TEX>$6.22\;{\mu}g/ml$</TEX>), PA도 10시간째(<TEX>$1.57\;{\mu}g/ml$</TEX>)에 각각 최대혈중농도에 도달하였다. PA의 조피볼락 혈중 내 흡수정도는 <TEX>$23{\pm}1.5^{\circ}C$</TEX>보다 <TEX>$13{\pm}1.5^{\circ}C$</TEX>에서 매우 높게 나타났으며, NA는 수온에 따른 차이가 크지 않았다. NA 및 PA의 조피볼락 혈중 내 소실정도는 <TEX>$13{\pm}1.5^{\circ}C$</TEX> 보다 <TEX>$23{\pm}1.5^{\circ}C$</TEX>에서 두드러지게 빨랐다. NA 및 PA는 one-compartment model로 해석(WinNonlin program)하여 약물동태학적 변수(parameter)를 조사하였다. <TEX>$23{\pm}1.5^{\circ}C$</TEX>의 경우, 혈장농도-시간곡선하 면적(AUC)은 NA, PA가 각각 161.25 및 <TEX>$41.57\;{\mu}g{\cdot}h/ml$</TEX>, 반감기(<TEX>$T_{1/2}$</TEX>)는 0.15 h 및 0.58 h, 혈중최고농도의 도달 시간(<TEX>$T_{max}$</TEX>)은 12.29 및 8.24 h, 혈중최고농도(<TEX>$C_{max}$</TEX>)는 3.85 및 <TEX>$0.21\;{\mu}g/ml$</TEX>로 계산되었다. <TEX>$13{\pm}1.5^{\circ}C$</TEX>의 경우, AUC는 NA, PA가 각각 134.42 및 <TEX>$40.36\;{\mu}g{\cdot}h/ml$</TEX>, <TEX>$T_{1/2}$</TEX>은 0.18 h 및 0.59 h, <TEX>$T_{max}$</TEX>는 10.67 및 21.15 h, <TEX>$C_{max}$</TEX>는 8.91 및 <TEX>$3.09\;{\mu}g/ml$</TEX>로 계산되었다. Effects of temperature (<TEX>$13{\pm}1.5^{\circ}C$</TEX>, <TEX>$23{\pm}1.5^{\circ}C$</TEX>) on the pharmacokinetic properties of nalidixic acid (NA) and piromidic acid (PA) were studied after oral administration to cultured black rockfish, Sebastes schlegeli. Serum concentrations of NA and PA were determined using HPLC-UV detector after a single dosage of 60 mg/kg body weight. At <TEX>$23{\pm}1.5^{\circ}C$</TEX>, the peak serum concentrations of NA and PA, which attained at 24 h post-dose, were 5.87 and <TEX>$0.43\;{\mu}g/ml$</TEX>, respectively. At <TEX>$13{\pm}1.5^{\circ}C$</TEX>, the peak serum concentrations of NA and PA, which attained at 10 h post-dose, were 6.22 and <TEX>$1.57\;{\mu}g/ml$</TEX>, respectively. Better absorption of PA was noted at <TEX>$13{\pm}1.5^{\circ}C$</TEX> compared to <TEX>$23{\pm}1.5^{\circ}C$</TEX>. However, absorption of NA was not affected significantly by temperature. The elimination of NA and PA from serum of black rockfish was considerably faster at <TEX>$23{\pm}1.5^{\circ}C$</TEX> than at <TEX>$13{\pm}1.5^{\circ}C$</TEX>. The kinetic profile of absorption, distribution and elimination of NA and PA in serum were analyzed by fitting to a one compartment model, with WinNonlin program. The AUC, <TEX>$T_{1/2}$</TEX>, <TEX>$T_{max}$</TEX> and <TEX>$C_{max}$</TEX>, respectively, were: <TEX>$161.25\;{\mu}g{\cdot}h/ml$</TEX>, 0.15 h, 12.29 h and <TEX>$8.91\;{\mu}g</TEX>/ml$ at <TEX>$23{\pm}1.5^{\circ}C$</TEX>, and <TEX>$134.12{\mu}g{\cdot}h/ml$</TEX>, 0.18 h, 8.79 h and <TEX>$5.00\;{\mu}g/ml$</TEX> at <TEX>$13{\pm}1.5^{\circ}C$</TEX> with NA; <TEX>$41.57\;{\mu}g{\cdot}h/ml$</TEX>, 0.58 h, 8.24 h and <TEX>$0.21\;{\mu}g/ml$</TEX> at <TEX>$23{\pm}1.5^{\circ}C$</TEX>, and <TEX>$40.36\;{\mu}g{\cdot}h/ml$</TEX>, 0.59 h, 5.04 h and <TEX>$1.20\;{\mu}g/ml$</TEX> at <TEX>$13{\pm}1.5^{\circ}C$</TEX> with PA.

Trace analysis of quinolone and fluoroquinolone antibiotics from wastewaters by liquid chromatography–electrospray tandem mass spectrometry

A sensitive liquid chromatography-electrospray tandem mass spectrometry method, combined with solid-phase extraction and a weak cation exchange cartridge cleanup, was established for twenty quinolone and fluoroquinolone antibiotics (pipemidic acid, flerofloxacin, ofloxacin, pefloxacin, enoxacin, norfloxacin, ciprofloxacin, danofloxacin, enrofloxacin, lomefloxacin, difloxacin, sarafloxacin, gatifloxacin, sparfloxacin, moxifloxacin, cinoxacin, oxolinic acid, nalidixic acid, flumequine, and piromidic acid) in influent, effluent, and river waters. For the various water matrices considered, the overall recoveries were from 64% to 127% except for piromidic acid (27-33%), and no obvious matrix effect was observed. The method detection limits for the twenty target antibiotics in the influent, effluent, and surface water samples were 1.6-50 ng/L, 0.6-50 ng/L, and 0.8-50 ng/L, respectively. This method was applied to analyze residual quinolone and fluoroquinolone antibiotics in wastewater and surface water samples from Beijing, China. Eight antibiotics (12 (pipemidic acid)-1208 ng/L (ofloxacin)) were detected in wastewater, and seven (1.3 (lomefloxacin)-535 ng/L (ofloxacin)) were detected in surface water samples. Gatifloxacin, a 4th generation fluoroquinolone antibiotic, was detected for the first time in influent (111 ng/L), effluent (56 ng/L), and river water (16-42 ng/L).

Separation of fifteen quinolones by high performance liquid chromatography: Application to pharmaceuticals and ofloxacin determination in urine

A simple chromatographic method is described for assaying 15 quinolones and fluoroquinolones (pipemidic acid, marbofloxacin, enoxacin, ofloxacin, norfloxacin, ciprofloxacin, danofloxacin, lomefloxacin, enrofloxacin, sarafloxacin, difloxacin, oxolinic acid, nalidixic acid, flumequine and piromidic acid), in urine and pharmaceutical samples. The determination was achieved by LC using an RP C18 analytical column. A mobile phase composed of mixtures of methanol-ACN-10 mM citrate buffer at pH 3.5 and 10 mM citrate buffer at pH 4.5, delivered under an optimum gradient program, at a flow rate of 1.5 mL/min, allows to accomplish the chromatographic separation in 26 min. For detection, diode-array UV-Vis at 280 nm and fluorescence detection set at excitation wavelength/emission wavelength: 280/450, 280/ 495, 280/405 and 320/360 nm were used. Detection and quantification limits were between 0.3-18 and 0.8-61 ng/mL, respectively. The method was validated in terms of interday (n = 6) and intraday (n = 6) precision and accuracy. The procedure was successfully applied to the analysis of human and veterinary pharmaceuticals. Also, ofloxacin was determined in human urine samples belonging to a patient undergoing treatment with this active principle, among others.

Multiresidue Determination of Quinolone and Fluoroquinolone Antibiotics in Fish and Shrimp by Liquid Chromatography/Tandem Mass Spectrometry

A multiresidue method was developed to measure low levels of 8 fluoroquinolones (norfloxacin, ofloxacin, danofloxacin, ciprofloxacin, desethylene ciprofloxacin, enrofloxacin, sarafloxacin, and difloxacin) and 4 quinolones (oxolinic acid, flumequine, nalidixic acid, and piromidic acid). Method detection limits range from 0.1 ng/g for quinolones to 0.4 ng/g for fluoroquinolones. Average recoveries range from 57 to 96%, depending on analyte and commodity; relative standard deviations are all less than 18%. The drugs are extracted from tissues using a mixture of ethanol and 1% acetic acid, diluted in aqueous HCI, and defatted by extraction with hexane. The compounds are further isolated using cation-exchange solid-phase extraction and measured using liquid chromatography with electrospray tandem mass spectrometry detection. The method has been evaluated and applied to the analysis of salmon, trout, and shrimp. Detectable residues were observed in 10 out of 73 samples, at concentrations ranging from 0.28 to 16 ng/g.

In-situ enumeration and probing of pyrene-degrading soil bacteria

Inferences about which microorganisms degrade polycyclic aromatic hydrocarbons in contaminated soils have largely been obtained using culture-based techniques, despite the low percentage of microorganisms in soil that are believed to be culturable. We used a substrate-responsive direct viable count method to identify and quantify potential polycyclic aromatic hydrocarbon-degrading bacteria in a soil containing petroleum wastes. Bacteria were extracted and their response to substrates determined in the presence of DNA gyrase inhibitors, which cause viable and active cells to elongate. When yeast extract, a widely used carbon source, was added as a growth substrate, together with nalidixic acid, piromidic acid and ciprofloxacin, a significant increase in elongated cells to 47%, 37% and 22%, respectively, was observed within 24 h. With pyrene as the main substrate, 10 mg L(-1) of nalidixic acid or piromidic acid caused 18-22% and 8-12%, respectively, of the cells to elongate within 24 h; whereas the effect of 0.5 mg L(-1) ciprofloxacin was not significant until 53 h later. Enlarged cells were identified and enumerated by fluorescent in situ hybridization, using Alpha-, Beta- and Gammaproteobacteria, and domain Bacteria-specific probes. The Bacteria-specific probe detected 35-71% of the total microorganisms detected by the DNA-binding dye 4,6-diamidino-2-phenylindole. Initially, 44%, 13% and 5% of the total bacteria in the soil extract were Alpha-, Beta- and Gammaproteobacteria, respectively. Without pyrene or a gyrase inhibitor, these subgroups decreased to 30% of the total population but were predominant with piromidic acid or unchanged with ciprofloxacin when pyrene was the main substrate. The proportion of elongated Alpha- and Betaproteobacteria (potential pyrene degraders) increased significantly (P<0.05). This approach links phylogenetic information with physiological function in situ without the conventional cultivation of bacteria and can be used to probe and enumerate degradative groups at even a finer level of discrimination.

Genotoxic potential of quinolone antimicrobials in the in vitro comet assay and micronucleus test

The purpose of this study was to examine the genotoxicity of quinolone antimicrobials. We investigated the genotoxic potential of eight quinolones, namely nalidixic acid (NA), pipemidic acid (PPA), oxolinic acid (OA), piromidic acid (PA), enoxacin (ENX), ofloxacin (OFLX), norfloxacin (NFLX) and ciprofloxacin (CPFX), by the in vitro alkaline single-cell gel electrophoresis (comet) assay at pH > 13. WTK-1 cells (mutant p53) were treated with each of the eight quinolones at 62.5–1000 μg/mL for 2, 4 and 20 h. NFLX and CPFX significantly induced DNA damage concentration-dependently after 4 and 20 h treatment, but this damage was recoverable. On the other hand, DNA was not damaged in the cells treated with six other quinolones. In the cells treated with NFLX and CPFX for 20 h, DNA migration was compared by the comet assay at pH 10, 12.1 and >13. The comet assay both at pH 12.1 and >13 showed increased DNA migration, but there was no positive response in the comet assay at pH 10. In the in vitro micronucleus (MN) test, WTK-1 cells were treated with each of four quinolones (NA, PPA, NFLX and CPFX) at 15.63–125 μg/mL for 20 h. NFLX significantly increased MNs in the cells, but no changes were noted in the cells treated with three other quinolones. These results suggest that NFLX and CPFX induced DNA single strand breaks (SSBs), and that NFLX-induced SSBs resulted in chromosome aberrations.

Membrane Preconcentration of Nalidixic and Piromidic Acids and Simultaneous Determination Using a Synchronous Solid‐Phase Phosphorescence Approach

A preconcentration membrane for the retention and phosphorimetric determination of pharmaceuticals nalidixic and piromidic acids is proposed. The membrane has a circular zone of plasticized PVC adhered to the surface of a polyester strip, which constitutes the preconcentration area where the analytes are retained by absorption from a solution that contains it. Measurement of intrinsic phosphorescence in the synchronous phosphorescence spectra performed at 281/371 nm for nalidixic acid and 331/471 nm for piromidic acid make the resolution of the mixture possible. The applicable concentration range, detection limit, and precision (as relative standard deviation) are from 0.3 to 3.0 mg · l−1, 0.08 mg · l−1, and 5.1% for nalidixic acid and from 0.03 to 0.15 mg · l−1, 0.002 mg · l−1, and 5.2% for piromidic acid. The method was applied to the determination of residues of both pharmaceuticals in cow milk and human urine.

In Vitro Activities of 25 Quinolones and Fluoroquinolones against Liver and Blood Stage Plasmodium spp

The in vitro activities of 25 quinolones and fluoroquinolones against erythrocytic stages of Plasmodium falciparum and against liver stages of Plasmodium yoelii yoelii and P. falciparum were studied. All compounds were inhibitory for chloroquine-sensitive and chloroquine-resistant P. falciparum grown in red blood cells. This inhibitory effect increased with prolonged incubation and according to the logarithm of the drug concentration. Grepafloxacin, trovafloxacin, and ciprofloxacin were the most effective drugs, with 50% inhibitory concentrations of <10 micro g/ml against both strains. Only grepafloxacin, piromidic acid, and trovafloxacin had an inhibitory effect against hepatic stages of P. falciparum and P. yoelii yoelii; this effect combined reductions of the numbers and the sizes of schizonts in treated cultures. Thus, quinolones have a potential for treatment or prevention of malaria through their unique antiparasitic effect against erythrocytic and hepatic stages of Plasmodium.

Square wave adsorptive stripping voltammetric determination of piromidic acid. Application in urine

A simple procedure for the determination of piromidic acid by square wave adsorptive stripping voltammetry (SW-AdSV) at a hanging mercury drop electrode has been developed. The variables affecting to accumulation process such as concentration of perchloric acid, accumulation potential and accumulation time have been optimised (0.025 mol L −1, −0.25 V and 140 s, respectively) by using response surface methodology. A linear relationship between concentration of piromidic acid and peak intensity has been found in the range 2.22×10 −9 to 3.33×10 −8 mol L −1. The detection limit (1.65×10 −9 mol L −1) has been calculated by the method proposed by Clayton et al. so that protection against both false positive and false negative errors is assured. The procedure was successfully applied to determine piromidic acid in spiked urine samples. The obtained recovery values were in the range 97.3–103.3% at different levels of concentration of piromidic acid.

Determination of quinolones and fluoroquinolones in fish tissue and seafood by high-performance liquid chromatography with electrospray ionisation tandem mass spectrometric detection.

A reversed-phase high-performance liquid chromatographic method with tandem mass-spectrometric detection was developed and validated for the simultaneous analysis of eight quinolones and fluoroquinolones (oxolinic acid, flumequine, piromidic acid, enrofloxacin, ciprofloxacin, danofloxacin, sarafloxacin and orbifloxacin) in trout tissue, prawns and abalone. The analytes were extracted from homogenised tissue using acetonitrile and the extracts subjected to an automated two-stage solid-phase extraction process involving polymeric reversed-phase and anion-exchange cartridges. Good recoveries were obtained for all analytes and the limit of quantification was 5 microg/kg (10 microg/kg for ciprofloxacin). The limit of detection was 1-3 microg/kg, depending on the analyte and matrix. Confirmation of the identity of a residue was achieved by further tandem mass-spectrometric analysis. A procedure for estimating the uncertainty associated with the measurement is presented.