Iranian Journal of Pharmacology and Therapeutics Razi Institute for Drug Research (RIDR) of Iran University of Medical Sciences and Health Services (IUMS) ISSN: 1735-2657 Vol. 4, Num. 2, 2005, pp. 91-94

Iranian Journal of Pharmacology & Therapeutics, Vol. 4, No. 2, 2005, pp. 91-94

Pharmacokinetic Studies Enrofloxacin in Yak after Intramuscular Administration

SANJIB KHARGHARIA, CHANDANA CHOUDHURY BARUA, PRITAM MOHAN and MOHAN BHATTACHARYA

Department of Pharmacology and Toxicology, College of Veterinary Science, Khanapara, Assam, India (S.K., C.C.B., P.M.); National Research Centre on Yak, Dirang, Arunachal Pradesh, India (M.B.).

Address correspondence to: Dr Sanjib Khargharia, Deptart-ment of Pharmacology and Toxicology, College of Veterinary Science, Khanapara, Guwahati-781022, India. E-mail: kharukharu@sify.com

Received July 27, 2005; Revised October 20, 2005; Accepted December 26, 2005

Code Number: pt05020

ABSTRACT

The pharmacokinetic studies of enrofloxacin were investigated in yak (Bos grunniens L.) after administra-tion of enrofloxacin by intramuscular route at 5 mg·kg^-1 body weight. Blood samples were collected from the jugular vein at predetermined time intervals after drug administration. Plasma was separated and ana-lysed for enrofloxacin by reverse-phase high performance liquid chromatography. Various pharmacoki-netic parameters were calculated using a non-compartmental model. The elimination half-life (t_½), area under plasma concentration-time curve (AUC), area under the moment curve (AUMC),mean residence time (MRT), apparent volume of distribution (V_d), total body clearance (Cl_B) and apparent first-order elimi-nation rate constant (K) of enrofloxacin were 2.79±0.60 h, 5.35±1.29 μg·h·mL^-1 , 21.50±8.60 μg·h²·mL^-1 , 4.02±0.87 h, 3.76±0.83 L·kg^-1 , 935.09 ± 236.45 mL·h^-1 ·kg^-1 and 0.25±0.07 h^-1 , respectively after the ad-ministration of enrofloxacin. Because of faster elimination time, excellent tissue penetration, shorter half- life, low MRT after intramuscular administration of enrofloxacin at a dose rate of 5 mg·kg^-1 , it is recom-mended for use in yaks at therapeutic dose.

Keywords: Enrofloxacin, Yak, Pharmacokinetics, Intramuscular

Enrofloxacin, a fluoroquinolone antimicrobial is ap-proved exclusively for veterinary use, has a broad spec-trum of antibacterial activity with MIC value ranging from 0.008 to 0.06 µg/mL. It has widespread distribu-tion to most tissues and body fluids with a potential therapeutic application in many types of infections. Hence, it is used effectively in the treatment of septice-mia, respiratory tract, urinary tract, skin, soft tissues, bone and joint infections etc. Yak (Bos grunniens L.), being the most ecologically sustainable animal re-sources of Indian Himalayas and is also mainstay for highlanders, provides basic needs in terms of meat, milk, hair, wool and most needful transport in hilly ter-rain. For control of disease in various animals pharma-cokinetic properties of enrofloxacin have been investi-gated in cattle [ 1 ], horses [ 2 ] and pigs [ 3 ]. The dosage regimens of enrofloxacin have been worked out for sheep, dog, and goat also, but such data are inadequate to warrant its effective clinical use in yak. In the light of the above reports, it was thought worthwhile to study pharmacokinetics of enrofloxacin to rationalize the dose and frequency of administration of enrofloxacin in yak (Bos grunniens L.).

MATERIALS AND METHODS

Experimental Animals

The study was conducted in six male yaks (Bos grunniens L) reared in the National Research Centre on Yak, Nykmadung Farm , Dirang, Arunachal Pradesh,. The animals weighed between 270-330 kg and 2_½-3½ years of age. The animals were examined clinically to evaluate health status and to rule out the possibility of any diseases. They were housed in the animal shed with concrete floor and were maintained on green fodder, dry grass and concentrate. Water was provided ad libitum.

Drugs and Chemicals

Technical grade and pure enrofloxacin, which was used as external standard in HPLC assay, were gener-ously gifted by Ranbaxy India Ltd, Ghaziabad. Other reagents for HPLC were procured from EMerck and Heparin from Sigma chemicals.

Experimental Design

The study was conducted in six clinically healthy male yaks. Pharmacokinetics studies of enrofloxacin was carried out after its single intramuscular administra-tion of 5 mg.kg^-1 body weight enrofloxacin. Blood sam-ples (2-3 mL) were collected by jugular vein puncture into heparinised tubes. The samples prior to and after administration of drug were collected at various time intervals up to 96 hr following enrofloxacin administra-tion. Plasma was harvested by centrifugation at 3000 rpm for 15 min and stored at –5°C till analysis for enro-floxacin.

Estimation of Enrofloxacin

For quantitative determination of enrofloxacin in plasma, the HPLC method of Teja–Isavadharm et al., [ 4 ] was followed with some modifications.

Instrumentation and Chromatographic Condi-tions. Plasma analysis was performed on a HPLC sys-tem (Perkin-Elmer, series 200, USA) fitted with a qua-ternary pump, Diode Array Detector, Auto Sampler and a Data Station. A 5 μm Hypersil BDS C18 (250×4.6 mm) HPLC column was used. The mobile phase con-sisted of 0.1 M phosphoric acid adjusted to pH 2.5 with a solution of 45% potassium hydroxide and acetonitrile mixed in a ratio of 70:30% (v/v). The flow rate of mo-bile phase was 1.2 mL. min^-1 and the eluent was moni-tored with Diode Array Detector adjusted wavelength at 290 nm. The chromatograms were integrated on a data station.

Sample Processing. Plasma samples were subjected to liquid phase extraction. To 1 mL of plasma, 1 mL of methanol was added, vortexed for 20 sec, then placed on ice for 15 min and centrifuged at 3500 rpm for 10 min. 750 µL supernatant was transferred to a test tube, 6 mL of dichloromethane was added to it and vortexed for 20 sec, followed by centrifugation at 1500 rpm for 10 min. The organic phase was transferred to a clean glass tube and evaporated to dryness at 40ºC. The residue was then reconstituted in mobile phase (500 µL) and 10 µL is injected into the column.

Blank plasma was spiked with standard parent com-pound at three different concentrations ranging from 2.5 to 10 µg. Plasma containing drug was extracted by liq-uid extraction procedure as described above and 10 µL was injected into the HPLC column to enable calibra-tion curve to be prepared. The plasma concentrations of enrofloxacin in samples were determined by comparing the detector response for the drug in the sample with the corresponding peak area in the standard mixture.

Analysis. Plasma concentrations versus time data of enrofloxacin obtained during the study were utilized for calculating various pharmacokinetic parameters using non-compartmental method of analysis [ 5 , 6 ]. Differ-ences between respective means for Pharmacokinetic parameters were evaluated using Student’s‘t’ test. P values <0.01 were considered to be statistically signifi-cant.

RESULTS

The mean plasma concentrations at different time in-terval after single dose of intramuscular administration at 5 mg·kg^-1 body weights in yak have been incorpo-rated in Fig 1 and presented in Table 1 .

Evaluation of the results on plasma level of enro-floxacin was adequately described by non-compartmental method. Mean pharmacokinetic parame-ters such as the area under plasma concentration-time curve (AUC), area under the moment curve (AUMC),mean maximum plasma concentration (C_max), mean time of maximum plasma concentration (t_max),mean resi-dence time (MRT), elimination half-life (t_½), apparent volume of distribution (V_d), total body clearance (Cl_B) and apparent first-order elimination rate constant (K) of enrofloxacin after i.madministration of enrofloxacin are presented in Table 2 . Following administration of the drug, the mean pharmacokinetic parameters of enro-floxacin calculated were t_½, 2.79±0.60 h; AUC, 5.35±1.29 μg·h·mL^-1 ; AUMC, 21.50±8.60 μg·h².mL^-1 ; MRT, 4.02±0.87 h; K, 0.25±0.07 h^-1 ; Cl_B, 935.09±236.45 mL·h^-1 ·kg^-1 and V_d, 3.76±0.83 L kg-1 respectively.

DISCUSSION

In the present study, enrofloxacin was administered intramuscularly at the dose of 5 mg·kg-1. The same dose has been used for determining the pharmacokinetics of enrofloxacin after intramuscular administration in horses [ 2 ], rabbits [ 7 ], and buffalo bulls [ 8 ].

Following the intramuscular administration, the therapeutic concentration was achieved within 0.04 h (2.5 min) and detected up to 24 h. The plasma concen-tration of enrofloxacin at 0.04 h (2.5 min) post admini-stration was 0.1667 ± 0.0143 μg·mL^-1 .

The maximum concentration of enrofloxacin was 2.1807±0.5712 μg·mL^-1 recorded at 0.75 hr (45 min) following intramuscular administration to healthy yaks. The peak plasma level obtained in this study was lower to C_max of 2.8±0.289 μg·mL^-1 reported by Rao [ 9 ] after intramuscular administration of enrofloxacin at the same dose rate in goat. But Kaartinen et al [ 1 ] reported much lower C_max of 0.7 μg·mL^-1 after i.m. administration of enrofloxacin in lactating cows.

The mean time of maximum plasma concentration (t_max) obtained in the present study was 0.75±0.14 h (45±8.66 min). This was comparable to t_max of 0.875±0.55 h obtained in rabbit after i.m. administration of enrofloxacin reported by Broome et al [ 10 ].

The mean overall elimination (K) rate constant was 0.2487±0.067 h^-1 in the present study was quite similar to the value (0.283±0.024 h–1) obtained after intramus-cular administration of enrofloxacin in goats [ 9 ].

The elimination half-life (t_½) of enrofloxacin after intramuscular administration obtained in the present study was 2.7866±0.6006 h which was higher than that of goats (t_½, 1.396 h) reported by Rao [ 9 ] but much lower than that of horse (9.9 h) after intramuscular ad-ministration of enrofloxacin [ 2 ].

The area under the plasma concentration time-curve (AUC) of enrofloxacin after its intramuscular admini-stration was 5.3471±1.286 μg·h·mL^-1 . The AUC ob-tained in this study was comparable to that reported for goats (7.516 μg·h·mL^-1 ) [ 9 ] after i.m. administration of enrofloxacin at 5 mg·kg^-1 body weight and cattle (7.1 μg·h·mL^-1) after subcutaneous administration [ 11 ].

Apparent volume of distribution (Vd_area) obtained in the present study was 3.7599±0.8297 L·kg^-1. This was comparatively higher than the Vd_area (1.19 L·kg^-1) re-ported by Gracia et al [ 12 ] in calves and 1.52 L·kg^-1 in goat [ 9 ]. In the present study the high Vd_area reflected excellent tissue penetration of enrofloxacin.

The bioavailability of enrofloxacin after intramuscu-lar administration was compared by the ratio of AUC_i.m . / AUC_i.v . In this study, a mean bioavailability of 93.29 % was obtained which indicated almost complete absorption of the drug after intramuscular administra-tion. However, Rao [ 9 ] reported higher bioavailability of 119 % in goat after intramuscular administration. But the present value is higher than that of the values of sheep (85%) after i.m. administration as reported by [ 13 ] Mengozzi et al.

The present disposition study reveals that for main-taining MIC of 0.06 µg/mL in plasma of yak, enroflox-acin should be given in the dose of 5 mg/kg bodyweight at 10 hours intervals by i.m. route.

REFERENCES

1. Kaartinen L, Salonene M, Alli L, Pyorala S. Pharmacokinetics of enrofloxacin after single intravenous, intramuscular and subcu-taneous injections in lactating cows. J Vet Pharmacol Therap 1995;9:254-63.
2. Kaartinen L, Pam S, Pyorola S. Pharmacokinetics of enroflox-acin in horses after single intravenous and intramuscular admini-stration. Equine Vet J 1997;29:378-81.
3. Zeng Z, Geng Q, Zeng Z L, Feng QH. Pharmacokinetics and bioavailability of enrofloxacin in pigs. Chin J Vet Sci 1996;19:606-12.
4. Teja Isvadharm, P Keratithakul, D Wattl, G Webster HK, Ed-stein MD. Measurement of Ciprofloxacin in human plasma, whole blood and erythrocytes by high performance liquid chro-matography. Therap Drug Monitoring 1991;13:263-7.
5. Goodman Gilman A, Rall TW, Nies AS, Taylor P. The Pharma-cological Basis of Therapeutics. 8th ed. New York: McGraw-Hill; 1995. p. 3-32.
6. Singh B. Non-compartmental pharmacokinetics analysis of plasma level data through statistical moment approach. A work-sheet instance. ICAR Short Course on ‘Recent approaches in clinical pharmacokinetics and therapeutic monitoring of drugs. 1999.
7. Cabanes A, Margarita Josh M, Garcia A, Franscissa P. Pharma-cokinetics of enrofloxacin after intravenous and intramuscular injection in rabbits. Are J Vet Res 1992;53:2090-3.
8. Verma HK, Pangwakar GR, Chaudhary RK, Srivastava AK. Pharmacokinetics and dosage regimen of enrofloxacin in buffalo bulls after intramuscular administration. Vet Res Commun 1999;23:501-5.
9. Rao GS. Pharmacokinetic studies of enrofloxacin in normal and febrile goats. [dissertation]. Izatnagar: Indian Veterinary Re-search Institute Deemed University.; 1999.
10. Broome LR, Brooks LD, Babish GI., Copeland DD , Conzelman GM. Pharmacokinetics of enrofloxacin in rabbits. Am J Vet Res 1991;52:1835-41.
11. Stegemann M, Wollen TS, Ewert KM, Terhune TN, Copeland DD. Plasma pharmacokinetics of enrofloxacin administration to cattle at a dose of 7.5 mg/kg. J Vet Pharmacol Therap 1997;20 Suppl 1:22-3.
12. Gracia OH, Errecalde C, Prieto G, Luders C, Pulles I, Bereco-chea C, Fernandez M. Pharmacokinetic of enrofloxacin in calves. Archivos De Veternaria 1996;28:107-11.
13. Mengozi G, Intorre L, Bertine S, Sodani. Pharmacokinetics of enrofloxacin and its metabolite, ciprofloxacin after intravenous and intramuscular administration in sheep. Am J Vet Res 1996;57:1040-104.

Copyright © 2005 by Razi Institute for Drug Research (RIDR)

The following images related to this document are available:

Photo images