Chiral aspects of metabolism of antiinflammatory drug flobufen in human hepatocytes

The metabolism of the nonsteroidal antiinflammatory drug flobufen, 4-(2',4'-difluorobiphenyl-4-yl)-2-methyl-4-oxobutanoic acid, was studied in primary cultures of human hepatocytes prepared by two-step collagenase perfusion of livers from four donors. Racemic flobufen or its individual enantiomers, R-(+)- and S-(-)-flobufen were used as substrates. Aliquots of culture medium were collected during 24-h incubation. The time-dependent disappearance of flobufen enantiomers and the formation of metabolites (stereoisomers of dihydroflobufen (DHF)) in hepatocytes were measured by chiral HPLC. The reduction of flobufen in human hepatocytes was stereoselective ((+)-R-flobufen was preferentially metabolized) and stereospecific ((2R;4S)-DHF and (2S;4S)-DHF stereoisomers were mostly formed). Although the structure of flobufen is different from the profens (2-arylpropionates), flobufen undergoes chiral inversion in human hepatocytes. The inversion of R-(+)-flobufen to S-(-)-flobufen predominates. The individual DHF stereoisomers were incubated in hepatocyte cultures and their biotransformation studied. The unidirectional chiral inversion of (2S;4S)-DHF to (2R;4S)-DHF and (2R;4R)-DHF to (2S;4R)-DHF was observed. Stereoselective oxidation of the DHFs to flobufen was also detected. Thus, flobufen metabolism in primary cultures of human hepatocytes is much more complicated (via chiral inversion and DHF re-oxidation) than was presumed from a preliminary achiral point of view.     

Inversion of (R)- to (S)-ketoprofen in eight animal species

The (R)-enantiomer of the NSAID ketoprofen was administered orally at 20 mg/kg to a series of 8 animal species. In all species, a highly significant degree of inversion occurred after 1 h which varied from 27% (gerbil) to 73% (dog) and persisted or increased in plasma samples obtained 3 h after drug administration. Although the (R)-enantiomer was inactive as an inhibitor of cyclooxygenase, the analgesic effects of that isomer was almost the same as the (S)-isomer in animal analgesic assays, following oral administration of the drugs to mice and rats. Taken together, the present results suggest that (R)-ketoprofen administered alone functioned primarily as a prodrug for (S)-ketoprofen under the experimental conditions of this study. © 1995 Wiley-Liss, Inc.     

Stereoselective pharmacokinetics and metabolism of flobufen in guinea pigs

Stereoselective aspects of pharmacokinetics and metabolism of a chiral nonsteroidal antiinflammatory drug, flobufen, 4-(2', 4'-difluorobiphenyl-4-yl)-2-methyl-4-oxobutanoic acid, were studied in male guinea pigs after p.o. administration of racemic flobufen (rac-flobufen) at a dose of 10 mg/kg. Blood samples were collected at intervals over 16 h after the administration of rac-flobufen for the quantification of flobufen enantiomers and their respective metabolites in plasma by chiral high-performance liquid chromatography (HPLC). Compartmental pharmacokinetic analysis was used to determine pharmacokinetic parameters of R- and S-flobufen. The plasma concentrations of the S- and R-enantiomers differed significantly during the experimental period. The S/R-enantiomeric ratio in 7plasma reached a maximum value of 10.1 at 240 min postdose. The oral clearance value of R-flobufen was five times higher than S-flobufen. The other pharmacokinetic parameters (K(e), T(1/2), V(SS)/F, MRT) of the enantiomers also differed substantially. All four stereoisomers of the dihydrometabolite of flobufen were detected in plasma with varying concentrations. Metabolite 17203 [4-(2,4-difluorophenyl)-phenylacetic acid] exhibited a relatively longer residence time compared to that noted for the enantiomers of the parent compound. Pharmacokinetics of the flobufen enantiomers were stereoselective in guinea pigs. The metabolism of flobufen was complex. However, metabolite 17203 seemed to be the main metabolite of flobufen that may be responsible for its relatively long-lasting antiphlogistic and immunomodulatory effects.     

The effect of the enantiomers of ibuprofen and flurbiprofen on the beta-oxidation of palmitate in the rat.

The effects of the enantiomers of ibuprofen (0.25 and 0.50 mmol/kg b.w.) and flurbiprofen (0.01, 0.03, and 0.06 mmol/kg b.w.) on the beta-oxidation of palmitate were investigated in the rat. The mean cumulative exhalation of 14CO2 after ip administration of [U-14C]palmitic acid was significantly reduced over 6 h by ibuprofen at the higher dose but not at the lower dose for either enantiomer. There was no difference between the enantiomers, the reduction over 6 h being 31.3 and 33.0% for (R)- and (S)-ibuprofen, respectively. There was also a significant inhibition of beta-oxidation by flurbiprofen at all 3 doses. Again, there was no stereoselectivity evident in this inhibition. Flurbiprofen was much more potent than ibuprofen in eliciting this effect, the 0.01mmol/kg dose giving a similar reduction in beta-oxidation as observed for the 0.50 mmol/kg dose of ibuprofen. The data support the hypothesis that inhibition of the in vivo beta-oxidation of palmitate by ibuprofen and flurbiprofen is primarily via a nonstereoselective noncoenzyme A-dependent mechanism.     

Formation of glycine conjugate and (-)-(R)-enantiomer from (+)-(S)-2-phenylpropionic acid suggesting the formation of the CoA thioester intermediate of (+)-(S)-enantiomer in dogs.

It has been proposed that the chiral inversion of the 2-arylpropionic acids is due to the stereospecific formation of the (-)-R-profenyl-CoA thioesters which are putative intermediates in the inversion. Accordingly, amino acid conjugation, for which the CoA thioesters are obligate intermediates, should be restricted to those optical forms which give rise to the (-)-R-profenyl-CoA, i.e., the racemates and the (-)-(R)-isomers. We have examined this problem in dogs with respect to 2-phenylpropionic acid(2-PPA). Regardless of the optical configuration of 2-phenylpropionic acid administered, the glycine conjugate was the major urinary metabolite and this was shown to be exclusively the (+)-(S)-enantiomer by chiral HPLC. Both (-)-(R)- and (+)-(S)-2-phenylpropionic acid were present in plasma after the administration of either antipode, and further evidence of the chiral inversion of both enantiomers was provided by the presence of some 25% of the opposite enantiomer in the free 2-phenylpropionic acid and its glucuronide excreted in urine after administration of (-)-(R)- and (+)-(S)-2-phenylpropionic acid. The (+)-(S)-enantiomer underwent chiral inversion to the (-)-(R)-antipode when incubated with dog hepatocytes. These data suggests that both enantiomers of 2-phenylpropionic acid are substrates for canine hepatic acyl CoA ligase(s) and thus undergo chiral inversion, but that the CoA thioester of only (+)-(S)-2-phenylpropionic acid is a substrate for the glycine N-acyl transferase. These studies are presently being extended to the structure and species specificity of the reverse inversion and amino acid conjugation of profen NSAIDs.     

Stereoselective disposition and chiral inversion of KE-298, a new antirheumatic drug,in rats

The present study was an attempt to elucidate the relationship between stereoselective pharmacokinetics and protein binding of KE-298 and its active metabolites, deacetyl-KE-298 (M-1) and S-methyl-KE-298 (M-2). Metabolic chiral inversion was also investigated. The levels of unchanged KE-298 in plasma after oral administration of (+)-(S)-KE-298 to rats were lower than those of (−)-(R)-KE-298, whereas the levels of M-1 and M-2 after administration of (+)-(S)-KE-298 were higher than after (−)-(R)-KE-298. In vitro, rat plasma protein binding of (+)-(S)-KE-298 was lower than that of (−)-(R)-KE-298. In contrast, the binding of (+)-(S)-M-1 and (+)-(S)-M-2 was higher than that of (−)-(R)-M-1 and (−)-(R)-M-2. Displacement studies revealed that the (+)-(S) and (−)-(R)-enantiomers of KE-298 and their metabolites bound to the warfarin binding site on rat serum albumin. These results suggest that the stereoselective plasma levels in KE-298 and its metabolites were closely related to enantiomeric differences in protein binding, attributed to quantitative differences in binding to albumin rather than to the different binding sites. Unidirectional chiral inversion was detected after oral administration of either (−)-(R)-KE-298 or (−)-(R)-M-2 to rats both yielding (+)-(S)-M-2. Chirality 9:22–28, 1997 © 1997 Wiley-Liss, Inc.     

Factors affecting aggression in a captive flock of Chilean flamingos (Phoenicopterus chilensis)

The influence of pair bond status, age and sex on aggression rates in a flock of 84 captive Chilean flamingos at Zoo Atlanta was examined. Analysis showed no difference between aggression rates of male and female flamingos, but adult flamingos had higher rates of aggression than juveniles. There were also significant differences in aggression depending on pair bond status (single, same‐sex pair, male–female pair or group). Bonded birds were significantly more aggressive than single birds, which is consistent with the concept that unpaired birds are not breeding and do not need to protect pair bonds or eggs. Birds in typical pair bonds (male–female) and atypical pair bonds (same‐sex pairs or groups) exhibited similar rates of aggression. These results contribute to the existing body of research on aggression in captive flamingos. Zoo Biol 30:59–64, 2011. © 2010 Wiley‐Liss, Inc.     

Cuticular lipid diversification in Lasiommata megera and Lasiommata paramegaera: the influence of species, sex, and population (Lepidoptera: Nymphalidae)

Numerous studies have investigated the presence and the effectiveness of volatile pheromones in Lepidoptera. Conversely, very few studies have focused on the composition and the perception of the relatively low volatile components of cuticular mixtures. Yet, cuticular lipids are implied in the recognition processes of several solitary and social insects. In the present study, the cuticular signatures of the satyrid butterflies Lasiommata megera and Lasiommata paramegaera were examined by gas chromatography. General linear model and discriminant analyses on chemical data clearly revealed large differences between sexes, which showed the same diversification pattern in both species. Moreover, a strong diversification between the two species was found, as were differences among populations. These results represent a first step in demonstrating the communicative function of cuticular compounds in the L. megera / paramegaera complex. Moreover, the discrimination among different species and populations on the basis of cuticular mixtures could represent a platform for studying chemotaxonomy and chemical biogeography in butterflies, as already found in several other insect groups.  © 2007 The Linnean Society of London, Biological Journal of the Linnean Society , 2007, 91 , 703–710.     

Sex differences in the social behavior of wild spider monkeys (Ateles geoffroyi yucatanensis)

Competition for resources and the need for cooperation are reported to affect patterns of social interactions and thus the quality of social relationships in primates. Relationships may be described as high quality when both individuals behave in a way that benefits their partner. We investigated the distribution of a wide range of social behaviors across sex partner combinations of adult spider monkeys with specific reference to contested resources. Data were collected from two communities of wild spider monkeys (Ateles geoffroyi yucatanensis) in the Otoch Ma'ax Yetel Kooh reserve in Yucatan, Mexico. Affiliative behavior was exchanged most frequently between males, and as male-male aggression was rare, male-male social relationships were characterized as high quality. Female-female social relationships were best described as low quality as females showed no preference to be in proximity with other females and female-female affiliative behavior was rare. Relationships between the sexes generally were characterized by high rates of female-directed male aggression, although additional investigation into the effects of kinship and female reproductive state on male-female relationships is required before further conclusions can be drawn. Dyadic and coalitionary female-female aggression was significantly higher than expected in a feeding context, confirming that female spider monkeys primarily compete for access to food. Male-male aggression did not increase significantly when in the presence of females, but males embraced at higher rates when in mixed-sex subgroups. As embraces serve to reduce the likelihood of aggression during tense situations, high rates of male-male embraces in mixed-sex subgroups may mitigate male conflicts over access to females.     

Leaders of progressions in wild mixed-species troops of saddleback (Saguinus fuscicollis) and mustached tamarins (S. mystax), with emphasis on color vision and sex

Leadership of travel progression is an important aspect of group living. It is widely believed that trichromacy evolved to facilitate the detection and selection of fruit in the dappled light of a forest. Further, it has been proposed that in New World primate species, which typically contain a range of color vision phenotypes, at least one female in a group will be trichromatic (i.e., having three types of visual pigment, in contrast to the two types of pigment found in dichromatic individuals) and will lead the group to fruiting trees. We examine progression leadership within two wild mixed-species troops of saddleback (Saguinus fuscicollis) and mustached (Saguinus mystax) tamarins over a complete year. As whole units, the mixed-species troops were most frequently led by a mustached tamarin. This is the first time that mixed-species group leadership and individual leadership have been quantified in these tamarin species. In terms of single-species intragroup leadership, neither the visual status (dichromatic or trichromatic) nor the sex of individuals had a consistent effect across species. Saddleback tamarin groups were led by males more frequently than females, while evidence suggests that mustached tamarins may be female-led. The notion that all groups contain at least one trichromatic female that leads the troop to feeding trees was not supported.     

Enantioselective HPLC determination of E-6087, a new COX-2 inhibitor, in human plasma: Validation and pharmacokinetic application

E-6087 is a nonsteroidal anti-inflammatory compound that selectively inhibits cyclooxygenase-2. Because E-6087 has a chiral center, this compound is a racemic mixture of two stereoisomers, (+)-(R)-E-6087 (E-6231) and (-)-(S)-E-6087 (E-6232). A normal-phase liquid-chromatographic method for the enantioselective determination of E-6087 in human plasma was developed and validated. The samples were extracted using solid-phase extraction cartridges containing C(18) sorbent, and the extracts were redissolved in absolute ethanol and injected into the chromatographic system. The enantiomeric separation was achieved on a chiral stationary-phase column of derivatized amylose, and the enantiomers were quantified by fluorescence detection. The method was validated for drug concentrations ranging from 5 to 400 ng/ml for both enantiomers. No peaks interfering with the quantification of enantiomers were observed. The limit of quantification was 5 ng/ml, with precision expressed as a coefficient of variation lower than 10.6% and accuracy expressed as relative error lower than 12.2%. The utility of this method was demonstrated by analysis of plasma samples from healthy volunteers given an oral dose of rac-E-6087. Peak plasma levels of E-6231 were higher than levels obtained for E-6232. Results were consistent with those obtained with a conventional reversed-phase method used for determination of the racemic compound.     

Stereoselective protein binding of tetrahydropalmatine enantiomers in human plasma,HSA, and AGP,but not in rat plasma

Tetrahydropalmatine (THP) is one of the active alkaloid ingredients of Rhizoma Corydalis. THP has a chiral center, and the stereoselective pharmacokinetics and tissue distribution have been reported. The aim of the present article is to study the stereoselective protein binding of THP using equilibrium dialysis followed by HPLC‐UV analysis. The results showed that THP stereoselectively binds to human serum albumin (HSA), α₁‐acid glycoprotein (AGP), and proteins in human plasma. The fraction binding of (+)‐THP was significantly higher than that of (?)‐THP, whereas such stereoselectivity was not found in rat plasma. The affinity of HSA and AGP to (+)‐THP, expressed as nK_A, were 9.0 × 10³ M^?1 and 2.34 × 10⁵ M^?1, respectively, which were notablely higher than to (?)‐THP, with the nK_A of 3.4 × 10³ M^?1 and 1.44 × 10⁵ M^?1, respectively. The binding site of HSA for (?)‐THP was Site I, whereas for (+)‐THP was both Site I and Site II. The F1/S variants of AGP were proved to be the key variants (?)‐ and (+)‐THP binding to both. Finally, the AGP binding drugs, such as mifepristone, were demonstrated to reduce the fraction binding of (?)‐ and (+)‐THP with pure AGP (1 mg/ml) but did not affect the fraction binding of both (?)‐ and (+)‐THP with proteins in human plasma. It can be concluded that protein binding of THP is species dependent and stereoselective, both HSA and AGP contribute to the stereoselective binding to THP enatiomers, and AGP binding drugs may not cause the drug–drug interaction on THP in healthy human plasma. Chirality, 2010. © 2009 Wiley‐Liss, Inc.     

Sex differences in the stone tool-use behavior of a wild population of burmese long-tailed macaques (Macaca fascicularis aurea)

We investigated sex differences in how Burmese long-tailed macaques (Macaca fascicularis aurea) used stone tools to open shelled food items along the shores of two islands in Laemson National Park, Thailand. Over a 2-week period in December 2009, we collected scan and focal samples on macaques when they were visible along the shores and mangroves. We found females used stones more often while feeding and used smaller tools than males. Females also processed sessile oysters more than males, whereas males processed unattached foods more than females. It was unclear which sex was overall more proficient at stone tool use, but males did perform significantly better at opening unattached food items with large pounding stones. Females also struck food items more times during tool-use bouts and at a faster rate, but no significant difference was found in average tool-use bout duration. Males processed foods slightly faster within a tool-use bout, but we were unable to detect a significant difference in the rate of food processing while foraging with tools. In summary females chipped open sessile oysters with an axing technique more than males, while males used larger stones to pound open unattached shelled food more often than females. Despite using pounding more than females, males also regularly utilized the axing technique on sessile oysters. Our results are the first assessment of sex differences in macaque stone tool use, providing a basis for comparison with tool use in other primates, and to nonfunctional forms of stone use in other macaques.     

Maternal antibodies in a wild altricial bird: effects on offspring immunity, growth and survival

1. In many animals immunity is not fully developed until adulthood but the young still need protection against various sets of pathogens. Thus, bird nestlings are highly dependent on antibodies received from their mother (in the eggs) during their rapid early growth period. The relationship between maternal immunity and the development of neonates' own immunity has been poorly studied. 2. It has been suggested that immune function plays an important part in mediating resource competition between different life-history traits, e.g. growth and reproduction. Maternal investment of antibodies has potentially permanent effects on offspring phenotype. Thus, the trade-offs between the immune function and other important life-history traits in the offspring will also affect the fitness of the mother. 3. Our supplemental feeding experiment in the magpie Pica pica indicates that the immunoglobulin levels of offspring at hatching are dependent on a mother's nutritional condition. In addition, the amount of maternal immunoglobulins transferred to offspring increases along the laying order within a nest. 4. We also found that at the age of 8-10 days the immunoglobulin production of the offspring has already begun. Furthermore, the maternal immunoglobulin levels of the offspring at hatching were positively related to their immunoglobulin levels on day 10. 5. Maternal immunoglobulins did not significantly affect offspring growth, but there was a negative relationship between self-produced immunoglobulins and growth over the first 10 days, indicating a trade-off between these traits. Nestlings' weight, however, had a positive relationship with immunoglobulin production suggesting that the observed trade-off between growth and immunoglobulin production is due to catch-up growth of nestlings with a low hatching weight. We found that within nests nestlings with higher maternal antibody levels had higher survival rate until day 20, but between nests there was an opposite relationship. 6. Evidently, there is a trade-off, in magpies, between maternal resources, immune function and growth, shaping the evolution of maternal investment in offspring immunity.     

Indirect chiral separation and analyses in human biological fluids of the stereoisomers of a thienothiopyran-2-sulfonamide (TRUSOPT), a novel carbonic anhydrase inhibitor with two chiral centers in the molecule.

The indirect chiral separation of the four stereoisomers (1)-(4) of a novel carbonic anhydrase inhibitor with two chiral centers in the molecule is reported. The method is based on chemical derivatization of the secondary amino group of the inhibitor with chiral isocyanate, formation of diastereomeric urea derivatives, each with three chiral centers in the molecule, and their separation under nonchiral HPLC conditions. The attempts to separate racemic mixture (1) + (2) from its diastereomeric counterpart (3) + (4) under nonchiral conditions, and to separate enantiomers (1) and (2) directly on a chiral stationary phase (CSP) are also reported. The indirect method was utilized for the assessment of an in vivo inversion of configuration at either one or both chiral centers of the molecule of (1). Analyses of selected whole blood and urine samples from human subjects after multiple bilateral topical ocular dosing with (1) did not reveal the presence of any of the three possible stereoisomers (2)-(4) of (1) indicating that the inversion of configuration at neither one nor two chiral centers of (1) occurs in vivo.     

High-speed microchip electrophoresis method for the separation of (R,S)-naproxen

In this research, a capillary electrophoretic method for the fast enantiomeric resolution of (R,S)-naproxen was investigated. Method development involved variation of applied potential, buffer concentration, buffer pH, and cyclodextrin concentration. The optimum electrophoretic separation conditions were 110 mM sodium acetate run buffer (pH 6.0), 30 mM methyl-beta-cyclodextrin, 20% (v/v) acetonitrile, 25 degrees C. The total length of capillary was 48 cm, (50 microm I.D.) with ultra violet (UV) detection at 232 nm. Using these conditions, the number of theoretical plates was close to one million (896,000/m). The possibility of achieving a fast chiral separation of (R,S)-naproxen on a microchip of 2.5 cm in length was investigated. Complete enantiomeric resolution of naproxen was achieved in less than 1 min, on this microchip platform, with linear imaging UV detection. This system had the advantage of real-time separation monitoring, so that enantiomeric resolution could be visually observed, and high-speed chiral analysis was realized. The microchip electrophoresis (MCE) separation was compared with the capillary electrophoresis (CE) separation with regards to speed, efficiency, separation platform, and precision. This work highlights the potential of CE and MCE in future chiral separations.     

Biology-oriented drug synthesis (BIODS), in vitro urease inhibitory activity,and in silico study of S-naproxen derivatives

Current study is based on the biology-oriented drug synthesis (BIODS) of S-naproxen (NSAID) derivatives and the evaluation of their urease inhibitory potential. In this regard, a variety of S-naproxen derivatives 2–39 including hydrazide 1, Schiff bases 2–21, aroyl substituted hydrazides 22–24, sulfohydrazides 25–34, 2-mercapto oxadiazole 35, phenacyl substituted 2-mercapto oxadiazoles 36–39 were synthesized under the umbrella of BIODS by simple chemical transformation of its pharmacophoric carboxylic group. Compounds 1–39 were evaluated for in vitro urease inhibitory activity and most of them showed good to moderate inhibitory potential in the range of IC₅₀ = 14.01 ± 0.23–76.43 ± 0.8 µM as compared to standard acetohydroxamic acid (IC₅₀ = 27.0 ± 0.5 µM). Limited structure-activity relationship (SAR) was established in order to rationalize the participation of varying groups (R) in the inhibitory potential of compounds. Molecular docking study on all active compounds was also carried out to decipher the interactions detail of the ligand with the receptors of active site of enzyme.     

Dental and general health in a population of wild ring-tailed lemurs: a life history approach

Data are presented on dental and general health for seven groups of wild ring-tailed lemurs, Lemur catta, from the Beza Mahafaly Reserve, in southern Madagascar. As part of a study of population demography, adults were captured, collared, and tagged, and biometric measurements, dental casts, and analyses of dental and general health were made. Results indicate that patterns of dental health vary by individual, age, sex, and habitat. Prime adults show more dental attrition than young adults. Prime males living in more marginal habitats show greater mean attrition than those living in richer habitats. Dental damage, specifically to the toothcomb, indicates that mechanical stresses to this region may include the initial harvesting of foods, in addition to grooming. Males exhibit more evidence of past trauma, including scars and chipped teeth. These results indicate that environmental as well as social factors, such as female dominance, may lead to sex differences in health patterns among lemurs.     

(R)- and (S)-5-hydroxy-2-(dipropylamino)tetralin (5-OH DPAT): assessment of optical purities and dopaminergic activities

Racemic 5-hydroxy-2-(dipropylamino)tetralin (5-OH DPAT), a potent and selective dopamine (DA) D2-receptor agonist, was resolved into the enantiomers by a new method. The enantiomers of 5-OH DPAT were determined by chiral ion-pair chromatography using N-benzyloxycarbonylglycyl-L-proline as the counter ion. The enantiomeric purity of (R)-5-OH DPAT was found to be greater than 99.7%. The ability of the enantiomers to change the rat brain DOPA levels was evaluated in vivo. The results indicate that (R)-5-OH DPAT is a weakly potent DA D2-receptor antagonist.