Simultaneous determination of proline and pipemidic acid in human urine by capillary electrophoresis with electrochemiluminescence detection.

Pipemidic acid is extensively used in the treatment of Gram-negative urinary tract infections, and the contents of proline in human urine vary in association with chronic uremia. The simultaneous determination of pipemidic acid and proline in human urine is of significance for quality control of the dosage and clinical study. The coupling of Ru(bpy)(3)(2+)-based electrochemiluminescence detection with capillary electrophoresis was developed for the simultaneous determination of proline and pipemidic acid in human urine. Parameters related to the separation and detection were investigated and optimized. The standard curves were linear between 0.1 and 90 µg mL(-1) for proline and between 0.4 and 100 µg mL(-1) for pipemidic acid. Underoptimized conditions, the detection limits (3σ) were 0.02 µg mL(-1) for proline and 0.06 µg mL(-1) for pipemidic acid. Relative standard derivations for the electrochemiluminescence intensity and the migration time were 3.2 and 0.9% for proline and 3.7 and 1.2% for pipemidic acid, respectively. The developed method was successfully applied to determine proline and pipemidic acid in human urine. The result showed that the content and decreasing rates of proline in urine for male were higher than that for female, and the content and decreasing rate of pipemidic acid in urine for male and female were consistent, respectively.

A capillary zone electrophoretic method for simultaneous determination of seven drugs in pharmaceuticals and in human urine.

A simple, sensitive, and novel method has been developed and validated for the separation and simultaneous quantitation of seven structurally different drugs-pipemidic acid and ofloxacin quinolone antibiotics, pseudoephedrine decongestant, piroxicam anti-inflammatory, thiamin, pyridoxine, and cobalamin-in a mixture by capillary zone electrophoresis. Factors affecting the separation were pH, concentration of buffer, and applied voltage. Separation was carried out in < 9 min with a 50 mM sodium tetraborate buffer, pH 10, and an applied voltage of 30 kV in an uncoated silica capillary tube. The carrier electrolyte gave baseline separation with good resolution, reproducibility, and accuracy. Calibration plots were linear over at least three orders of magnitude of analyte concentrations, and the lower LODs were within the range of 1-5 microg/mL. Detection was performed by UV absorbance at 230 nm. The method was validated for the analysis of drugs in pharmaceutical preparations and in urine samples with RSD of 0.5-2.4% and recovery of > 99%.

Electrochemiluminescence determination of pipemidic acid using sulfite as energy transfer mediator.

An electrochemiluminescence (ECL) based on energy transfer from electro-generated triplet sulfur dioxide to pipemidic acid (PPA) was studied. A weak ECL from triplet sulfur dioxide (3)SO2 * was observed when sulfite was electrochemically oxidized in sulfuric acid solution on a Pt electrode. When PPA was present, the weak ECL was enhanced. The enhanced ECL was attributed to energy transfer from (3)SO2 * to PPA. Based on the enhanced ECL, a flow-injection (FIA) ECL method for the determination of PPA was proposed. The proposed method allowed the measurement of PPA over the range of 1.0x10(-7) to 2.0x10(-5)moll(-1). The detection limit was 3.9x10(-8)moll(-1), and the relative standard deviation for 1.0x10(-6)moll(-1) PPA (n=9) was 1.3%. This method was evaluated by the analysis of PPA in pharmaceutical preparations and urine samples.

Fluorimetric determination of pipemidinic acid using the europium chelate.

The sensitized luminescence of europium ion in the complexes with pipemidinic acid is investigated. It was shown that in the result of intramolecular energy transfer from ligand to lanthanide ion the luminescence intensity of the latter increases by 10(10) times. The luminescence properties of the complex were studied and the high sensitive luminescence method for the determination of pipemidinic acid has been developed.

[Study of the fluorescence of terbium-pipemidic acid system and the determination of pipemidic acid in urine and serum].

Intramolecular energy transfer occurs in the terbium complex with pipemidic acid (PPA) and the terbium(III) ion fluorescence can be sensitized by PPA. The fluorescence intensity of the Tb3+-PPA system is proportional to the amount of PPA. Accordingly, a fluorimetric method for the determination of PPA is proposed. The characteristics of the method include low background noise, high selectivity and long fluorescence lifetime. The method can be applied to the determination of PPA in urine and serum. The recovery is in the range of 94.6%-102%. Its detection limit and linear range for PPA are 3.6 ng/mL and 5.0 x 10(-8)-4.0 x 10(-6) mol/L, respectively.

Determination of the chemotherapeutic quinolonic and cinolonic derivatives in urine by high-performance liquid chromatography with ultraviolet and fluorescence detection in series.

An HPLC method with ultraviolet and fluorimetric detection has been established for the separation and determination of six quinolonic and cinolonic antibiotics. A Nova-Pak C18 column (150 x 3.9 mm) and a Waters 486 UV and a Waters 470 fluorescence detector have been used. The influence of variables such as mobile-phase composition and flow-rate, has been studied. An acetonitrile-aqueous solution of oxalic acid 4x10(-4) M (28:72, v/v) has been selected as optimum. The wavelength for the photometric detection of the six antibiotics was 265 nm. For the fluorimetric detection two pairs of excitation/emission wavelengths, 260/360 or 270/440 nm, were selected for the determination of nalidixic acid, 7-hydroxymethylnalidixic acid and oxolinic acid, and for the determination of pipemidic acid and cinoxacin, respectively. The analytical parameters and detection and quantification limits of the method have been determined. The proposed method has been applied for the determination of the six compounds in urine, applying different procedures depending on their concentration, the results being very acceptable.

Adsorptive stripping voltammetric determination of pipemidic acid in human urine.

The electrochemical behaviour of pipemidic acid (8-ethyl-5,8-dihydro-5-oxo-2-(1-piperazinyl)-pyrido[2,3-d]pyrimidine-6- carboxylic acid), a well known antimicrobial agent used for urinary infections, was investigated by linear-sweep, differential-pulse and square-wave voltammetry at a hanging mercury drop electrode. Two reduction processes were observed in Britton-Robinson buffers at acid pH, whereas only one or two processes were observed in alkaline solutions, dependent on the pH of the buffer employed. Adsorptive effects were used to accumulate the drug on to the electrode. The adsorbed species were measured voltammetrically by using a cathodic process appearing at -0.76 V in 0.1 M HCIO4. Linear calibration graphs were obtained in the range 2.5 x 10(-9)-2.0 x 10(-7) M. A simple procedure of extraction was employed for the determination of the drug in urine samples.

Antibacterial activity in human urine of fosfomycin trometamol in an in vitro model of the urinary bladder.

The urinary concentrations of fosfomycin trometamol, norfloxacin, pipemidic acid and cotrimoxazole were studied at various times after oral administration of drugs in healthy volunteers. Using the same urine, the bactericidal activity of four antimicrobial agents against Escherichia coli, Proteus mirabilis and Klebsiella pneumoniae in an in vitro model simulating the treatment of bacterial cystitis was also evaluated. The results obtained show that very high concentrations of the drugs were achieved in urine particularly after the oral administration of the fosfomycin trometamol. In the bladder model bactericidal activity of fosfomycin trometamol, norfloxacin and pipemidic acid were higher than that of cotrimoxazole; no resistant mutants to drugs were selected over a period of 24 h.

Antibacterial activity after a single-dose of norfloxacin, ofloxacin and pipemidic acid detected in urine of volunteers.

Six healthy volunteers received in a triple-crossover design a single oral dose of norfloxacin, ofloxacin or pipemidic acid. Urine samples were collected during the 24 h following the administration and were tested for inhibitory and bactericidal activity against selected strains of Enterobacteriaceae and Pseudomonas aeruginosa. Norfloxacin and ofloxacin inhibited the urinary growth of sensitive strains during the 24 h of sampling time at dilutions much higher than those generally considered satisfactory. Nalidixic acid was less effective and did not achieve bactericidal activity against Ps. aeruginosa over the interval of 12 to 24 h.

[Studies on the pharmacokinetics of pipemidic acid in renal insufficiency (author's transl)]. / Untersuchungen zur Pharmakokinetik der Pipemidsäure bei eingeschränkter Nierenfunktion.

After an oral single dose of 400 mg pipemidic acid administered to 7 patients with renal insufficiency of varying degrees of severity, serum concentrations were determined by microbiological and spectrofluorimetric methods. The results obtained revealed maximum values ranging between 1.8 and 7.2 micrograms/ml with a biological half-life of 5.7--16 h. Of some therapeutic significance was an increase in serum concentrations amounting to 2.6--12.8 micrograms/ml when pipemidic acid was administered at normal doses (2 X 400 mg/d) for 3 days. Under maintenance therapy maximum concentrations of microbiologically active substance in urine were found to range between 177 and 580 micrograms/ml. Even in severely impaired kidney function urine concentrations averaged 230 micrograms/ml, which indicates that therapeutically sufficient concentrations can be maintained by regular administration of normal doses at unaltered dosage intervals.

Metabolites of pipemidic acid in human urine.

1. The urine of men dosed orally with pipemidic acid, contained the metabolites acetylpipemidic, formylpipemidic and oxopipemidic acids together with unchanged pipemidic acid. 2. Each metabolite was equivalent to less than 2% of the unchanged pipemidic acid present in human urine. 3. All metabolites showed a similar antibacterial spectrum to pipemidic acid, but their potency was about 10 times lower than that of pipemidic acid. The acute toxicity of the metabolites in mice was as low as the parent compound. 4. These results indicate that the role of the metabolites in the clinical efficacy and toxicity of the drug is likely to be small.