Simultaneous Determination of Eight ß-Lactam Antibiotics, Amoxicillin, Cefazolin, Cefepime, Cefotaxime, Ceftazidime, Cloxacillin, Oxacillin, and Piperacillin, in Human Plasma by Using Ultra-High-Performance Liquid Chromatography with Ultraviolet Detection.

A simple and rapid ultra-high-performance liquid chromatography (UHPLC) method using UV detection was developed for the simultaneous determination of eight ß-lactam antibiotics in human plasma, including four penicillins, amoxicillin (AMX), cloxacillin (CLX), oxacillin (OXA), and piperacillin (PIP), and four cephalosporins, cefazolin (CFZ), cefepime (FEP), cefotaxime (CTX), and ceftazidime (CAZ). One hundred-microliter samples were spiked with thiopental as an internal standard, and proteins were precipitated by acetonitrile containing 0.1% formic acid. Separation was achieved on a pentafluorophenyl (PFP) column with a mobile phase composed of phosphoric acid (10 mM) and acetonitrile in gradient elution mode at a flow rate of 500 µl/min. Detection was performed at 230 nm for AMX, CLX, OXA, and PIP and 260 nm for CFZ, FEP, CTX, and CAZ. The total analysis time did not exceed 13 min. The method was found to be linear at concentrations ranging from 2 to 100 mg/liter for each compound, and all validation parameters fulfilled international requirements. Between- and within-run accuracy errors ranged from -5.2% to 11.4%, and precision was lower than 14.2%. This simple method requires small-volume samples and can easily be implemented in most clinical laboratories to promote the therapeutic drug monitoring of ß-lactam antibiotics. The simultaneous determination of several antibiotics considerably reduces the time to results for clinicians, which may improve treatment efficiency, especially in critically ill patients.

On-line clean-up and screening of oxacillin and cloxacillin in human urine and plasma with a weak ion exchange monolithic column.

A weak ion exchange monolithic column prepared by modifying the GMA-MAA-EDMA (glycidyl methacrylate-methacrylic acid-ethylene glycol dimethacrylate) monoliths with ethylenediamine was applied to remove matrix compounds in biological fluid. Using this monolithic column, on-line clean-up and screening of oxacillin and cloxacillin in human urine and plasma samples had been investigated. Chromatography was performed by reversed-phase HPLC on a C(18) column with ultraviolet detection at 225 nm. Results showed that the ion exchange monolithic column could be used for deproteinization and retaining oxacillin and cloxacillin in human urine and plasma, which provided a simple and fast method for assaying drugs in human urine and plasma.

Effect of pentobarbital anaesthesia on intestinal absorption and hepatic first-pass metabolism of oxacillin in rats, evaluated by portal-systemic concentration difference.

The effects of anaesthesia on intestinal drug absorption and hepatic first-pass metabolism in rats were investigated by observing the difference in the drug concentration between portal and systemic bloods. Oxacillin and pentobarbital were selected as a model drug and as an anaesthetic, respectively. Rats were divided into a conscious control group and an anaesthetized group. All rats were cannulated simultaneously in the portal vein and in the femoral artery, and oxacillin was orally administered after its intra-arterial injection (double dosing). For the anaesthetized group, pentobarbital was intrasubcutaneously administered twice, first before intra-arterial injection and again before oral administration of oxacillin. The arterial blood alone was sampled from the cannula in the femoral artery before oral administration, whereas the arterial and portal bloods were simultaneously sampled from both cannulated sites after oral administration. Oxacillin concentrations in plasma were assayed by HPLC. The anaesthesia increased the absolute bioavailability (F), the mean absorption time (MAT) and the hepatic recovery ratio (F(H)), but caused little change in the local absorption ratio into the portal system (Fa) and the total clearance (CL). The hepatic clearance (CL(H)) was significantly decreased, resulting in an apparent small change in CL-CL(H) which is considered to be renal clearance. By this method, it was shown directly that an increase in F due to pentobarbital anaesthesia was attributable to the significant increase in F(H). It is expected that the method is useful not only to evaluate the effect of anaesthesia on the first-pass effect, but also to assess the effect of co-administration of drugs on first-pass metabolism.

Effect of liver intoxication by carbon tetrachloride on hepatic local disposition of oxacillin using moment characteristics as index.

The effect of liver intoxication by carbon tetrachloride (CCl4) on the hepatic local disposition using oxacillin as a test drug and bovine serum albumin (BSA) as a reference substance was evaluated by single-pass bolus input method in the isolated perfused rat liver. Oxacillin and BSA were introduced into the liver from the portal vein, and the outflow concentration-time profiles of oxacillin and BSA from the rat liver into the hepatic vein were kinetically assessed by moment analysis. Liver damage was monitored by plasma SGOT, plasma SGPT, and bile flow rate. Hepatic recovery ratio FH of oxacillin increased from 50% to 80%, with an increase in liver intoxication by CCl4; whereas FH of BSA was naturally 100%. Mean transit time tH of oxacillin increased from 6 to 12 sec, with an increase in liver intoxication; whereas tH of BSA increased from 7 to 10 sec. The relative variance sigma 2/tH2 of oxacillin increased from 0.2 to 0.7, with an increase in liver intoxication; whereas sigma 2/tH2 of BSA took the value of approximately 0.4, independent of liver damage. In the relationship between the dispersion model and moments, it was shown that the blood space VB increased from 15 to 20%, the index for distribution (1 + k') of oxacillin from 1.0 to 1.5, the efficiency number RN of oxacillin decreased from 0.7 to 0.3 with the increase in liver damage, and the extent of eddy mixing was predicted to be unaffected by liver damage. The intensive increases in tH and sigma 2/tH2 of oxacillin with the liver intoxication by CCl4 were explained by increases in the extent of distribution and nonequilibrium distribution, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Septicemia por Staphylococcus aureus em crianças: tolerância bacteriana à vancomicina e poder bactericida do soro / Staphylococcus aureus septicemia in children: bacterial tolerance to vancomycin and serum bactericidal test

OBJETIVO. Analisar a sensibilidade e tolerância das cepas de Staphylococcus aureus isoladas de crianças com septicemia e avaliar o poder bactericida sérico na monitorizaçäo terapêutica desses casos. MÉTODOS. Foram estudados 17 casos de crianças com septicemia por Staphylococcus aureus internadas na Enfermaria de cuidados semi-Intensivos do Instituto da Criança do Hospital das Clínicas da Faculdade de Medicina da Universidade de Säo Paulo. Foram realizados testes de sensibilidade antimicrobiana pelo método de difusäo em disco e diluiçäo em tubo. Foram realizados 29 testes no pico e 23 no vale dos antibióticos utilizados, determinando o poder bactericida do soro. RESULTADOS. As cepas de Staphylococcus aureus de origem hospitalar mostraram resistência a quase todos os antibióticos, exceto vancomicina e pefloxacina. Observou-se fenômeno de tolerância em cinco (50 por cento) das cepas testadas para vancomicina, sendo que quatro apresentaram má evoluçäo clínica. Os testes para determinaçäo do poder bactericida sérico revelaram títulos no pico ò1/8 em 55,5 por cento das observaç 8es; neste grupo a evoluçäo clínica foi melhor. CONCLUSäO. As cepas de Staphylococcus aureus de origem hospitalar estudadas säo multirresistentes. O fenômeno de tolerância antimicrobiana, assim como o poder bactericida do soro em níveis baixos, pode estar associado a má resposta terapêutica. A valorizaçäo do PBS como critério de avaliaçäo terapêutica em infecçöes graves e o papel da tolerância do Staphylococcus aureus à vancomicina merecem maiores estudos

Single-dose antibiotic prophylaxis in maxillofacial surgery.

The clinical efficacy of short-term antimicrobial prophylaxis with either one shot of ceftriaxone (1 g) or a course of 3 injections of a fixed combination of mezlocillin (2 g) and oxacillin (1 g) administered over 24 h was studied in a prospective randomized clinical study of 100 patients undergoing elective maxillofacial surgery. Tissue and plasma concentrations of the antibiotics were determined by high-pressure liquid chromatography in 6 tumor surgery patients from each treatment group. Statistical analysis showed the treatment group to be comparable both demographically and with respect to the types of surgery performed and the durations of the procedures. Only 1 patient in each group developed a postoperative wound infection. It is concluded that 1 g ceftriaxone given 30 min preoperatively meets the pharmacokinetic requirements for perioperative antimicrobial prophylaxis in maxillofacial surgery.

Killing rate and serum bactericidal activity of oxacillin, rifampin and ciprofloxacin against Staphylococcus aureus.

The bacterial activity of oxacillin, rifampin and ciprofloxacin was examined at two concentrations by the serum bactericidal rate (SBR) technique against 5 strains of methicillin-susceptible Staphylococcus aureus. The activity of oxacillin plus ciprofloxacin and oxacillin plus rifampin was also determined for 5 strains of staphylococci examining in vitro the SBR and the serum bactericidal activity (SBA). We simulated SBR and SBA in vitro using pooled human serum containing know concentrations of antimicrobial agents. The rate at which the antibiotics kill S. aureus did not rise by increasing the concentrations over the MBC. The oxacillin-ciprofloxacin combination was indifferent when tested by the rate of killing, whereas an antagonistic interaction was frequently observed with the oxacillin-rifampin combination. The SBA was determined by two methods: the technique of the Mayo Clinic that uses Mueller-Hinton broth (MHB) as the diluent and the method of Stratton in which the diluent was prepared with MHB supplemented with Ca++ and Mg++ and combined with 50% pooled human serum. The activities of oxacillin and rifampin were decreased in the presence of human serum. These results were attributed to the high protein binding of these antibiotics. Rifampin in combination with oxacillin showed antagonism against S. aureus also by the SBA method. The inhibitory activity of drugs in combination remained substantially the same as the single more active one.

Oxacillin and tobramycin serum levels during cardiopulmonary bypass.

Antibiotic prophylaxis in cardiac surgery is recommended to combat acquired infections caused by staphylococci and gram-negative bacilli. Prophylaxis seems effective provided blood levels are greater than minimal inhibitory concentrations (MIC). In this study, two doses of antibiotics were compared in 45 patients with normal renal function during cardiopulmonary bypass (CPB). All patients received 50 mg/kg of oxacillin. Group 1 (30 patients) also received 1 mg/kg of tobramycin, while group 2 (15 patients) received 2 mg/kg of tobramycin. Blood samples were taken after the administration of antibiotics, as well as at the onset and conclusion of CPB. Additional samples were taken before and after heparin injection before CPB, and from the arterial and venous cannulae of the bubble oxygenator during CPB. In both groups, oxacillin serum levels were constantly greater than MIC for susceptible bacteria. In group 1, tobramycin levels less than 2 micrograms/mL (MIC for most susceptible bacteria) occurred in four patients before CPB, in 14 patients at the onset of CPB, and in 19 patients at the conclusion of CPB. These low levels were not explained by heparin administration or absorption onto the CPB circuit, but were the result of hemodilution. In group 2, in which all the tobramycin levels were higher than 2 micrograms/mL, serum levels decreased from 9.9 +/- 3.4 (mean +/- SD) to 3.7 +/- 0.7 micrograms/mL throughout the procedure. Plasma creatinine did not change significantly in either group. It is concluded that in patients with normal renal function, doses as high as 50 mg/kg of oxacillin and 2 mg/kg of tobramycin may be necessary before CPB to provide adequate serum levels throughout CPB.

[In vivo bactericidal activity of an oxacillin-netilmicin combination against Staphylococcus aureus. Influence of the rhythm of netilmicin administration]. / Etude de l'activité bactéricide in vivo de l'association oxacilline-nétilmicine vis-à-vis de Staphylococcus aureus. Influence du rythme d'administration de la nétilmicine.

The bactericidal activity of two regimens of netilmicin (8 mg/kg/day) given intravenously once a day (od) or thrice daily (tid) both alone and in combination with oxacillin (200 mg/kg/day) was compared using a model of fibrin clots infected with a strain of Staphylococcus aureus (10(7) CFU/g) sensitive to methicillin and netilmicin (clinical isolate) and implanted subcutaneously in rabbit. This study shows that: 1) Netilmicin given alone as both single and divided doses results in early bacterial killing but does not exert a bactericidal effect after 24 hours because of a significant late increase of the number of bacterial. 2) The netilmicin-oxacillin combinations are more bactericidal at 1 h, 2 h and 24 h than oxacillin alone (P less than 0.001). 3) The oxacillin-netilmicin combination appears to be better for bacterial killing when netilmicin is given thrice daily (P less than 0.001). It is hard to draw a clinical inference from such an experimental study but it seems that 8-hour divided doses intervals should be better for administration of netilmicin than single daily dose during the acute period of staphylococcal infections.

[The concentration of oxacillin and ampicillin (Penstabil) in serum and mother's milk in puerperal mastitis]. / Untersuchungen zur Konzentration von Oxacillin und Ampicillin (Penstabil) in Serum und Muttermilch bei der Mastitis puerperalis.

In 23 women in childbed suffering from mastitis puerperalis the transport of oxacillin and ampicillin into the milk was investigated. Moreover the concentration of the antibiotics in the urine of babies nursed during therapy of their mothers with antibiotics was measured. After a regular taking (good compliance) the drug-concentrations in the serum and the milk of the mothers are higher than the minimal inhibiting concentrations necessary against the number of bacteria found in the milk. In the urine of suckling babies drug concentrations were differently high or undetectable as well. During the therapy of nursing mothers with antibiotics the milk should be misplaced.

Influence of technical factor variations on serum inhibition and bactericidal titers.

The influence of technical factor variations on serum bactericidal and serum inhibitory titers was studied by using Staphylococcus aureus clinical isolates versus oxacillin-spiked human serum. Parallel tests, both with and without the use of beta-lactamase in count plates to inactivate oxacillin carryover, were performed with a conventional macrodilution approach, a carefully controlled macrodilution procedure, and a standard microdilution method. Careful control of technical factor variations diminished the incidence of low serum bactericidal titers and decreased the dispersion of results, a finding corollary to the known influence of technical factor variations on the measurement of MBCs. The incorporation of beta-lactamase into count plates resulted in a shift of serum bactericidal titers to lower values. The microdilution method appeared to be least influenced by technical variations and, with the addition of beta-lactamase to count plates, provided the best results.

[Serum level of antibiotics in burn patients]. / Der Serumspiegel von Antibiotika bei Brandverletzten.

Antibiotic serum levels were recorded from 100 patients with burns. Antibiotic doses to ensure effective serum levels were found to be in excess of maximum values quoted as necessary by manufacturers, provided that serum creatinine was normal. These findings were in agreement with literature data. This was the first instance to check on serum levels, following oral administration of antibiotics. Differences between individual patients may be considerable, probably due to variable renal blood supply and other factors. Individual dosage with control of serum levels is, therefore, recommended. Specific dosage recommendation for patients with burns would be desirable for hospital practice.

Comparison of serum and coagulum antibiotic levels with respect to protected blood coagulum.

Serum and coagulum antibiotic levels have a parallel course. Coagulum antibiotic concentrations are more than ten times lower than serum concentrations. Therefore the optimal dosage of perioperative infusion should be high enough to establish steady state by means of sufficiently high serum levels. Antibiotics capable of attaining high serum levels and having a long half-life are best suited for the formation of effective antibiotic concentrations in blood coagulum. This is particularly true for cephalosporins and penicillins. Aminoglycoside antibiotics are not recommended for the formation of protected coagulum. "Ceftriaxon in the treatment of Severe bacterial infections".

Double-blind, placebo-controlled study of oxacillin combined with rifampin in the treatment of staphylococcal infections.

A total of 101 patients with proven Staphylococcus aureus infection were included in a double-blind, placebo-controlled study; this study compared oxacillin (12 g/day, intravenously) or vancomycin (2 g/day, intravenously) plus rifampin (1,200 mg/day, orally) with oxacillin or vancomycin plus placebo. We evaluated 65 patients. Of the patients tested, 33 received oxacillin plus rifampin (13 bacteremias), and 32 received oxacillin plus placebo (16 bacteremias). Clinical cure was achieved in 61% of the patients treated with oxacillin plus rifampin and in 56% of the patients treated with oxacillin plus placebo. Improvement was noted in 27 and 25%, respectively, and failure occurred in 9 and 18%, respectively. These differences were not statistically significant. Bacteriological failure occurred in 3 and 28%, respectively (P less than 0.05). None of the failures within the rifampin-treated group was associated with the emergence of a rifampin-resistant mutant. The rates of superinfection were similar in both groups. The geometric means of the serum bactericidal activity after 1, 6, and 11 h were, respectively, 22, 17, and 9 after treatment with oxacillin plus rifampin and 25, 3.4, and 2.3 after treatment with oxacillin plus placebo. It was suggested that the addition of rifampin to oxacillin or vancomycin might only be beneficial to severely ill patients.

[Serum antibacterial activity during antibiotic treatment in automated bacteriology]. / Etude de l'activité antibactérienne sérique au cours des traitements antibiotiques en automate de bactériologie.

The conventional serum-dilution bactericidal test used for monitoring antibiotic therapy in severely infected in patients requires 72 h. Use of an automaton would be expected to provide faster results. We tested the MS-2 diagnostic automaton which performs kinetic analysis of bacterial growth. A relation between exponential growth onset time and initial bacterial concentration of the inoculum was first determined using successive ratio 10 dilutions. Sixty-two serum samples from twenty-one patients with endocarditis or septicemia were examined. A selection of findings is presented. Results of the automatic method and serum-dilution bactericidal test are studied comparatively. From our data we define the major advantages of automation: only 0.65 ml of serum and 10 minutes are needed to perform the test and results are obtained in five to twelve hours according to the bacterial strain. This new technique could be integrated in the monitoring protocol of severe infections.

[Penetration of ampicillin and oxacillin into the tissues of rats with aseptic inflammation]. / Izuchenie proniknoveniia ampitsillina i oksatsillina v tkani krys s asepticheskim vospaleniem.

Penetration of 2 penicillins with different indices of serum protein binding in the tissues of rats with aseptic inflammation was studied. The pharmacokinetics of both penicillins in the blood serum (total preparation) did not differ. However, the level of the ampicillin free fraction was much higher than that of oxacillin. In the inflammation exudate and inflamed tissues, the maximum concentrations of free ampicillin and the area below its pharmacokinetic curve were also higher than those of oxacillin. The rate of elimination of both antibiotics from the exudate was the same. The period of half elimination of the drugs from the exudate and inflamed tissue was significantly higher than the period of their half elimination from the blood serum. High positive correlation between the ampicillin levels in the blood serum and inflamed and intact tissues was shown. As for oxacillin, positive correlation between its levels in the blood serum and inflamed tissue was observed.

Comparison of oral penicillinase-resistant penicillins: contrasts between agents and assays.

We observed the comparative serum levels and mean peak serum antistaphylococcal activity in eight fasting adults who received 500 mg each of dicloxacillin, cloxacillin, oxacillin, and nafcillin. Dicloxacillin achieved higher and more prolonged serum levels and greater peak serum antistaphylococcal titers than the other drugs studied. The higher degree of protein binding of dicloxacillin was reflected in a greater disparity between the peak antistaphylococcal activity observed when dilutions were done in serum compared to broth. The lesser protein-bound penicillins showed less disparity, but this effect was offset by the higher serum levels obtained by dicloxacillin. The higher protein binding of dicloxacillin did not prevent its having equal or superior antistaphylococcal activity in serum when the drugs were given in equal doses.

[Mezlocillin and oxacillin concentrations in the bile of the choledochus]. / Mezlocillin- und Oxacillin-Konzentrationen in der Galle des Ductus choledochus.

The purpose of this study was to determine the antibiotic concentrations in serum and bile one and two hours after the completion of a simultaneous short-term infusion of 4 g mezlocillin and 2 g oxacillin. Seven patients were studied. Bile was withdrawn from the bile duct by means of a Teflon catheter. The mean serum concentrations of mezlocillin were 130.4 mg/l and 60.8 mg/l one and two hours after the end of the infusion, respectively. The mean serum concentrations of oxacillin determined at the same times were 57.9 mg/l and 18.9 mg/l, respectively. The mean biliary levels of mezlocillin were 3215.6 mg/l and 2483.1 mg/l one and two hours after the end of the infusion, respectively. The corresponding mean biliary values for oxacillin were 497.3 mg/l and 205.5 mg/l.

Oxacillin for bacterial endophthalmitis: subconjunctival, intravenous, both, or neither?

We compared the intraocular concentrations of oxacillin given by continuous intravenous infusion, subconjunctival injection, or combined therapy in a rabbit model of Staphylococcus aureus endophthalmitis. At equilibrium during intravenous infusion, concentrations in the aqueous humor, cornea, and choroid-retina were 25% to 30% of the serum level; in contrast, vitreous levels were only 2% of the serum concentration. Subconjunctival injection produced extremely high levels in the cornea and aqueous and moderate concentrations in the choroid-retina; vitreous penetration remained poor (<1 microgram/ml). Combined therapy offered little advantage in terms of concentrations in the aqueous or cornea and had a modest effect on levels in the choroid-retina. Vitreous concentrations showed a striking relation to serum levels with all regimens, including subconjunctival ones; although this could have been fortuitous, it suggests an important role for the hematogenous route in this model. Only those modes of delivery producing serum concentrations greater than 50 micrograms/ml consistently resulted in vitreous levels greater than 0.4 microgram/ml. The optimal therapy of bacterial endophthalmitis may require direct intravitreal injection of antibiotic.