Indian Journal of Pharmacology Medknow Publications on behalf of Indian Pharmacological Society ISSN: 0253-7613 EISSN: 1998-3751 Vol. 36, Num. 2, 2004, pp. 80-83

Indian Journal of Pharmacology, Vol. 36, No. 2, April, 2004, pp. 80-83

Research Paper

Ofloxacin pharmacokinetics in saliva

Hemanth Kumar AK, Gurumurthy P

Department of Biochemistry, Tuberculosis Research Centre (ICMR), Chennai - 600031
Correspondence Address:Department of Biochemistry, Tuberculosis Research Centre (ICMR), Chennai - 600031 icmrtr@vsnl.com

Code Number: ph04030

ABSTRACT

INTRODUCTION

Ofloxacin is a fluorinated carboxy quinolone exhibiting a marked bactericidal effect by inhibiting DNA gyrase.[1],[2] In vitro studies on bactericidal activity against Mycobacterium tuberculosis have suggested that ofloxacin is likely to be the most useful drug in the early stages of treatment and in preventing the emergence of resistance to other drugs.[3] Its favorable pharmacokinetic features include good oral absorption and lack of metabolism resulting in less drug interactions.[4]

The determination of drug concentrations in the saliva has gained widespread acceptance in a variety of settings.[5] The estimation of drugs in the saliva has been employed for therapeutic drug monitoring and for calculation of pharmacokinetic variables.[6] The rational use of such determinations can provide knowledge of patient-specific pharmacokinetic parameters leading to improved therapy.[7]

Saliva can serve as an alternative body fluid for pharmacokinetic investigations. It can be collected with minimal patient discomfort and can be easily obtained on multiple occasions. It is particularly suitable for investigations in geriatrics and pediatrics. Pharmacokinetic studies have shown that ofloxacin penetrates into saliva and its concentration correlates well with serum levels.[8] However, the data available in the published studies are inconsistent and results reported for the ratio of saliva to serum concentrations show wide variations.

The present study was undertaken to evaluate the pharmacokinetics of ofloxacin in saliva when administered alone or in combination with other antituberculosis (TB) drugs, viz. rifampicin (R), isoniazid (H) and pyrazinamide (Z) and to study the ratio of the saliva to plasma concentration by measuring ofloxacin concentrations in simultaneous saliva and plasma samples in healthy volunteers.

MATERIAL AND METHODS

Assay for the stated contents of the drugs
Before starting the study, six tablets of ofloxacin (Dec-Pharma Ltd.), R (Asojsofcaps Pvt. Ltd.), H (Pfizer) and Z (Lupin Laboratories) were assayed on five different occasions to check for the quantity and the stated contents. The drugs employed in the study were drawn from the same batch.

Study design
The volunteers were investigated on four different occasions with an interval of at least one week between occasions. They were administered ofloxacin on one occasion and ofloxacin in combination with R, H and Z on other occasions, as shown in [Table - 1]. A partially balanced incomplete block design was adopted. Thus, the volunteers received either ofloxacin alone or in combination with R, H, Z and RHZ. After an overnight fast, the drugs were administered on an empty stomach after emptying the bladder. The dosages of the drugs administered to the volunteers were approximately 10 mg/kg body weight for R, 35 mg/kg for Z and the dosages of ofloxacin and H were 10 mg/kg for subjects weighing < 40 kg and 15 mg/kg for subjects weighing > 40 kg.

Sample collection
On each occasion, 2-3 ml of paired samples of saliva and heparinized blood were collected at 1, 2, 3, 6 and 8 h after oral administration of the drug(s). Plasma was separated and stored. The study participants were instructed to wash their oral cavity and a piece of unsweetened, unflavored chewing gum was used to stimulate salivary secretion. The saliva samples were frozen at -20°C. They were thawed the following day, centrifuged and the supernatant was separated. The saliva and plasma samples were stored at -20°C. By storing biological samples under these conditions, no detectable drug decomposition takes place.

Assay of ofloxacin
Ofloxacin concentrations in saliva and plasma were determined by the plate diffusion method employing a strain of E.coli as described elsewhere.[9] In brief, the organism seeded in the autoclaved nutrient agar was poured into sterile petriplates. Wells of 2 mm diameter were cut. Ofloxacin standards ranging from 0.25-8.0 mg/ml in pooled saliva and plasma and the drug in saliva and plasma samples were set up in quadruplicate. The concentrations of the drug were obtained from the regression line of the zone of inhibition on log concentrations of the standards. The assays were undertaken after coding the samples.

Pharmacokinetic analysis
Peak ofloxacin concentration (C_max) in saliva and plasma as well as the time at which peak concentration was attained (t_max) were obtained from the measured data. The elimination rate constant (Kel) was calculated from the terminal log-linear decline of concentrations in the measured fluid. Terminal elimination half-life (t_½) was calculated as 0.693/Kel. The areas under the concentration-time curve (AUC) (0-8 h) for O were calculated by the trapezoidal rule. Bioavailability of O for each drug combination was expressed as an index (B.I.) and calculated as the ratio of drug combination to that of O alone (control).

Data analysis was performed using SPSS (version 10.5) package. Differences among the treatment groups were determined using one-way ANOVA. P values <0.05 were considered significant.

RESULTS

Based on the salivary concentrations of ofloxacin at different time points, several pharmacokinetic variables were calculated [Table - 2]. The mean Cmax values of ofloxacin when administered alone and in combination with R, H, Z and RHZ were 4.88, 5.38, 4.36, 4.63 and 5.18 mg/ml respectively. There were no significant differences in the Cmax and the tmax of ofloxacin between the different groups.

The concentrations of ofloxacin in saliva were much lower than those in plasma. The Cmax values of ofloxacin in plasma when administered alone and in combination with R, H, Z and RHZ were 7.14, 6.98, 7.19, 6.06 and 6.38 mg/ml respectively. The mean saliva to plasma ratio of ofloxacin concentrations was calculated at all time points and it was around 0.6. The correlation coefficient between plasma and saliva concentration of ofloxacin was 0.94. The mean saliva to plasma (S/P) ratio of ofloxacin concentration when given alone and in combination was compared [Table - 2]. There was no significant difference between the groups. The bioavailability indices of ofloxacin in saliva and plasma calculated on the basis of Cmax and AUC (in comparison with various combinations) are shown in [Table - 3]. They are similar in all the groups.

DISCUSSION

Ofloxacin has proved to be a particularly valuable addition to the available anti-TB drugs with a MIC of 1.0 mg/ml against Mycobacterium tuberculosis. The effectiveness of ofloxacin against Mycobacterium tuberculosis was studied by Crowle et al[10] and their results reveal its clinical usefulness.

Using saliva instead of blood for pharmacokinetic investigations has obvious practical advantages. It is a painless, non-invasive procedure, hence suitable for the collection of multiple specimens. Variable results have been reported for the ratio of saliva to serum concentrations. Koizumi et al[11] reported a ratio of 1.0 between 2-8 h after administration of a 300 mg dose. Fujita et al[12] in their studies on patients with renal impairment had reported a ratio of 1:1 for saliva to serum concentrations. However, Warlich et al,[13] have reported a ratio of 0.6 and a close relation of levels in the saliva and serum (r= 0.99), which is consistent with our findings. The mean S/P ratio of ofloxacin was around 0.6 and this remained the same when ofloxacin was given in any combination. The S/P ratio is a possible measure of the amount of protein binding to the drug. There was a good correlation (0.94) with respect to drug concentrations.

The present study shows that several pharmacokinetic parameters can be calculated using salivary concentrations of ofloxacin as can be done with plasma. The estimation of ofloxacin levels in saliva seems to be useful in therapeutic drug monitoring and pharmacokinetic studies.

Based on the salivary levels, the present study compares the pharmacokinetic parameters of ofloxacin when given alone and in combination with other anti-TB drugs namely R, H and Z. The pharmacokinetic parameters such as C_max, t_max, AUC, and t_½ did not change when given in combination with other anti-TB drugs suggesting that the pharmacokinetics of ofloxacin do not get affected or altered when administered with the other anti-TB drugs.

When ofloxacin was administered in combination with three other drugs, namely R, H and Z, there were variations in the pharmacokinetic parameters calculated based on the salivary estimates. This could be due to the small sample size in this group (n=4) and interindividual differences in the absorption and metabolism of the drug.

The microbiological assay (MBA) employed for the estimation of ofloxacin in the saliva and plasma is very simple, specific, sensitive and reproducible, and does not require sophisticated equipment. This method was evaluated in samples wherein drug levels were also measured by HPLC according to the method of Immanuel et al.[14] A very good correlation (0.99) was observed suggesting either method may be useful for pharmacokinetic studies.

It can, therefore, be concluded that for bioavailability studies, invasive blood collection can be replaced by simple non-invasive saliva collection. Further investigations are required to determine whether the concentration of ofloxacin in saliva can act as a valid index after multiple dosing. In this regard, a larger population sampling is required before any general statements regarding the utility of salivary measurements of ofloxacin can be made.

ACKNOWLEDGEMENTS

The authors wish to thank the volunteers for their willingness to participate in this study, Mr. L. Sekar for the statistical help and Mr. B. Doraiswamy for secretarial assistance.

REFERENCES

1.	Neumann M. Clinical pharmacokinetics of the newer antibacterial 4- quinolones. Clin Pharmacokinet 1988;14:96-121.  Back to cited text no. 1
2.	Verho M, Malerczyk V, Dagrosa E, Korn A. The effect of food on the pharmacokinetics of ofloxacin. Curr Med Res Opin 1986;10:166-71.  Back to cited text no. 2  [PUBMED]
3.	Herbert D, Paramasivan CN, Venkatesan P, Kubendran G, Prabhakar R, Mitchison DA. Bactericidal action of ofloxacin, sulbactum-ampicillin, rifampin and isoniazid on logarithmic and stationary-phase cultures of Mycobacterium tuberculosis. Antimicrob Agents Chemother 1996;40:2296-9.  Back to cited text no. 3
4.	Yuk JH, Nightingale CH, Quintiliani R, Sweeney KR. Bioavailability and pharmacokinetics of ofloxacin in healthy volunteers. Antimicrob Agents Chemother 1991;35:384-6.  Back to cited text no. 4  [PUBMED]
5.	Drobitch RK, Svensson CK. Therapeutic drug monitoring in saliva: An update. Clin Pharmacokinet 1992;23:365-79.  Back to cited text no. 5  [PUBMED]
6.	Mucklow JC. The use of therapeutic drug monitoring. Ther Drug Monit 1982;4:229-47.  Back to cited text no. 6
7.	Spectar R, Park GD, Johnson GF, Vesell ES. Therapeutic drug monitoring. Clin Pharmacol Ther 1988;43:345-53.  Back to cited text no. 7
8.	Guay DR. The role of the fluoroquinolones. Pharmacotherapy 1992;7:71-85.  Back to cited text no. 8
9.	Gurumurthy P, Ramachandran G, Hemanth Kumar AK, Venkatesan P, Chandrasekaran V, Narayanan PR. Simple spectrofluorimetric and microbiological assay methods for the estimation of ofloxacin in biological fluids. Indian J Pharmacol 1998;30:263-6.  Back to cited text no. 9
10.	Crowle AJ, Elkins N, May MH. Effectiveness of ofloxacin against Mycobacterium tuberculosis and Mycobacterium avium and rifampin against Mycobacterium tuberculosis in cultured human macrophages. Am Rev Respir Dis 1988; 137:1141-6.  Back to cited text no. 10  [PUBMED]
11.	Koizumi F, Ohnishi A, Takemura H, Okubo S, Kagami T, Tanaka T. Effective monitoring of concentrations of ofloxacin in saliva of patients with chronic respiratory tract infections. Antimicrob Agents Chemother 1994;38:1140-3.  Back to cited text no. 11  [PUBMED]
12.	Fujita K, Matsuoka N, Takenaka I. Pharmacokinetics of ofloxacin-measurement of drug concentrations in saliva of patients with impaired renal function. Drugs 1995;49:312-3.  Back to cited text no. 12
13.	Warlich R, Korting HC, Schafer-Korting M, Mutschler E. Multiple-dose pharmacokinetics of ofloxacin in serum, saliva and skin blisterfluid of healthy volunteers. Antimicrob Agents Chemother 1990;34:78-81.  Back to cited text no. 13  [PUBMED]
14.	Immanuel C, Hemanth Kumar AK. Simple and rapid high-performance liquid chromatography method for the determination of ofloxacin concentrations in plasma and urine. J Chromatogr B 2001;760:91-5.  Back to cited text no. 14  [PUBMED]

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