Despite the wealth of experimental data on cocaine abuse, there are no published dose-response pharmacokinetic studies with bolus i.v. cocaine injection in the male rat. The present study examined the pharmacokinetics of arterial plasma concentrations of cocaine and metabolite profile [benzoylecgonine (BE), ecgonine methyl ester (EME), norcocaine (NC)] following a single i.v. injection of 0.5, 1.0, or 3.0 mg/kg cocaine. Male Sprague-Dawley rats (N = 25) were anesthetized and surgically instrumented with both jugular vein (drug administration) and carotid artery (blood withdrawal) catheters and allowed to recover for at least 24 h. Arterial plasma samples (200 microliters) were obtained at eight time points (0.5, 1.5, 2.5, 10, 20, 30 min) following i.v. bolus injection (15-s injection, 15-s flush) and analyzed by single ion monitoring using GC/MS. Nonlinear regression and noncompartmental pharmacokinetic analysis were employed. Mean +/- SEM peak plasma concentrations of cocaine occurred at 30 s in a dose-response manner (370 +/- 14,755 +/- 119,2553 +/- 898 ng/ml for 0.5, 1.0, and 3.0 mg/kg groups, respectively). T1/2 alpha was < 1 min for all groups, but inversely related to dose. T1/2 beta was independent of dose 13.3 +/- 1.6, 13.0 +/- 1.5, and 12.0 +/- 2.0 min for 0.5, 1.0, and 3.0 mg/kg groups, respectively). MRT (16.0, 15.9, 14.5 min), VdSS (3.3, 3.2, and 2.8 l/kg), and ClTOT (204, 201, and 195 ml/min/kg) also provided little evidence of dose-dependent effects. Although the metabolic profile of i.v. cocaine was similarly ordered for all dose groups (BE > EME > NC), a quantitative shift in metabolite profile was evident as a function of increasing dose. This metabolic shift, perhaps attributable to saturation of plasma and liver esterases, suggests that the recently reported pharmacodynamic effects positively correlated with i.v. cocaine dose are unlikely attributable to NC, a minor but pharmacologically active metabolite. In sum, the i.v. pharmacokinetic profile in rats is distinct from that observed via the SC, IP, and PO routes of administration and offers the potential to provide a reasonable clinically relevant rodent model.

The complete nucleotide sequence of the rhsA locus and selected portions of other members of the rhs multigene family of Escherichia coli K-12 have been determined. A definition of the limits of the rhsA and rhsC loci was established by comparing sequences from E. coli K-12 with sequences from an independent E. coli isolate whose DNA contains no homology to the rhs core. This comparison showed that rhsA comprises 8,249 base pairs (bp) in strain K-12 and that the Rhs°strain, instead, contains an unrelated 32-bp sequence.Similarly, the K-12 rhsC locus is 9.6 kilobases in length and a 10-bp sequence resides at its location in the Rhso Families of homologous but nonidentical genes present a number of special genetic questions. These questions include the possible specific roles of individual members, the degree to which members exchange heterologies through intrachromosomal recombination, and the effects of these exchanges on function. The presence of these multigene families also has important implications for chromosome rearrangement and evolution (17). However, except for the rrn operons encoding ribosomal RNA, multigene families are quite rare in Escherichia coli (19). We have recently reported an unusual and complex family, the rhs family, that is comparable to the rrn family in number, length of shared homology, and degree of sequence similarity. The rhs loci were originally detected through their action as rearrangement hot spots, providing homology for recA-dependent intrachromosomal recombination (4, 11). Consequently, the rhs loci were defined to include the homologous sequences shared by two or more of the respective loci. Four rhs loci of E. coli strain K-12 have been characterized extensively, and evidence for a fifth has been noted (4,11,20).A distinctive feature of the rhs loci is that they share a highly conserved 3.7-kilobase (kb) core sequence. The cores are generally flanked by dissimilar sequences, and for two of the loci, rhsA and rhsC, one or more partial core repetitions are present downstream from the intact core. Sequence comparison of the first 300 nucleotides of the rhs cores (20) revealed that the core homology begins precisely with a start codon initiating an open reading frame (ORF) and that the rhsA, rhsB, and rhsC cores are closely related, showing only 1 to 2% sequence divergence. By contrast, rhsD is 18% divergent from the others. A total of nine mismatches distinguish rhsA, rhsB, and rhsC through these 300 nucleotides, but none of the nine causes an amino acid substitution in the core ORFs. However, rhsD differs from the others by eight amino acids. This degree of divergence through predominantly neutral mutation indicates that the cores have been evolving independently for quite some time. Application of the mutation rate estimated for enteric bacteria by Ochman and Wilson (15) suggests that the rhsA and rhsC cores diverged on the order of 10 million years ago, and the extent of sequence divergence of rhsD would indicate that it radiated from the others on the order of 100 mill...

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Lidocaine is a local anaesthetic agent that is widely used in equine medicine. It is also an Association of Racing Commissioners International (ARCI) Class 2 foreign substance that may cause regulators to impose substantial penalties if residues are identified in post race urine samples. Therefore, an analytical/pharmacological database was developed for this drug. Using our abaxial sesamoid local anaesthetic model, the highest no-effect dose (HNED) for the local anaesthetic effect of lidocaine was determined to be 4 mg. Using enzyme-linked immunosorbent assay (ELISA) screening, administration of the HNED of lidocaine to eight horses yielded peak serum and urine concentrations of apparent lidocaine of 0.84 ng/mL at 30 min and 72.8 ng/mL at 60 min after injection, respectively. These concentrations of apparent lidocaine are readily detectable by routine ELISA screening tests (LIDOCAINE ELISA, Neogen, Lexington, KY). ELISA screening does not specifically identify lidocaine or its metabolites, which include 3-hydroxylidocaine, dimethylaniline, 4-hydroxydimethylaniline, monoethylglycinexylidine, 3-hydroxymonoethylglycinexylidine, and glycinexylidine. As 3-hydroxylidocaine is the major metabolite recovered from equine urine, it was synthesized, purified and characterized, and a quantitative mass spectrometric method was developed for 3-hydroxylidocaine as recovered from horse urine. Following subcutaneous (s.c.) injection of the HNED of lidocaine, the concentration of 3-hydroxylidocaine recovered from urine reached a peak of about 315 ng/mL at 1 h after administration. The mean pH of the 1 h post dosing urine samples was 7. 7, and there was no apparent effect of pH on the amount of 3-hydroxylidocaine recovered. Within the context of these experiments, the data suggests that recovery of less than 315 ng/mL of 3-hydroxylidocaine from a post race urine sample is unlikely to be associated with a recent local anaesthetic effect of lidocaine. Therefore these data may be of assistance to industry professionals in evaluating the significance of small concentrations of lidocaine or its metabolites in postrace urine samples. It should be noted that the quantitative data are based on analytical methods developed specifically for this study, and that methods used by other laboratories may yield different recoveries of urine 3-hydroxylidocaine.