Species limits and phylogeographic structure in two genera of solitary African mole-rats Georychus and Heliophobius.

African mole-rats (Bathyergidae) are an intensively studied family of subterranean rodents including three highly social and three solitary genera. Although their phylogenetic interrelations are clear, genetic diversity and the number of species within each genus is much less certain. Among the solitary genera, Heliophobius and Georychus were for a long time considered as monotypic, but molecular studies demonstrated strong phylogeographic structure within each genus and proposed that they represent complexes of cryptic species. The present study re-evaluates their internal genetic/phylogenetic structure using a combination of methodological approaches. We generated datasets of one mitochondrial and six specifically selected nuclear markers as well as of a large number of double digest restriction site associated (ddRAD) loci and then applied species delimitation analyses based on the multispecies coalescent model or clustering on co-ancestry matrices. The population structure was largely congruent across all analyses, but the methods differed in their resolution scale when determining distinct gene pools. While the multispecies coalescent model distinguished five Georychus and between eleven to thirteen Heliophobius gene pools in both Sanger sequenced and ddRAD loci, two clustering algorithms revealed significantly finer or coarser structure in ddRAD based co-ancestry matrices. Tens of clusters were distinguished by fineRADstructure and one (in Georychus) or two clusters (in Heliophobius) by Infomap. The divergence dating of the bathyergid phylogeny estimated that diversification within both genera coincided with the onset of the Pleistocene and was likely driven by repeated large-scale climatic changes. Based on this updated genetic evidence, we suggest recognizing one species of Georychus and two species of Heliophobius, corresponding to a northern and southern major lineage, separated by the Eastern Arc Mountains. Yet, the final taxonomic revision should await integrated evidence stemming from e.g. morphological, ecological, or behavioral datasets.

Chiral resolution of flobufen by high performance liquid chromatography and capillary electrophoresis.

The enantiomeric separation of an anti-inflammatory drug flobufen was performed on various chiral columns in liquid chromatography and by reversed phase chromatography and by capillary electrophoresis with beta-cyclodextrin as a chiral selector in the mobile phase and background electrolyte, respectively. The elution order of individual enantiomers is discussed with respect to the absolute chirality of (+/-)-flobufen determined by X-ray crystallography.

Quantitation of collagen types I, III and V in tissue slices by capillary electrophoresis after cyanogen bromide solubilization.

A method for the determination of the proportions of major fiber-forming collagens (types I, III and V) in soft connective tissue was elaborated. The method is based on the release of insoluble collagen by CNBr with subsequent separation of the arising peptides. For routine application the peptides are separated by capillary electrophoresis (50 mM phosphate pH 2.5, 15 kV, 50 degrees C, 70/60 cm x 70 microns I.D. capillary with UV detection at 200 nm). Quantitation of collagen type I can be done either on the basis of spiking the sample with a peptide mixture obtained from a known amount of collagen type I, or by spiking the sample with an equimolar mixture of the two peptides [alpha 1(I)CB2 and alpha 1(I)CB4] (constituting a fused peak) along with alpha 1(III)CB2 and alpha 1(V)CB1. Compared to the previously published methods the procedure is faster and does not require isolation of marker peptides by tedious chromatographic procedures in a preceding preparatory step. Good results are obtained within a wide range of run buffer concentrations and applied voltages; conversely, intensive cleaning of the capillary after every three runs is recommended with a new capillary after 20-30 runs.

Separation of common nucleotides, mono-, di- and triphosphates, by capillary electrophoresis.

A capillary electrophoretic procedure for the separation of eleven nucleotides, 5'-mono-, di- and triphosphates of adenosine, guanosine, cytidine and uridine, has been developed. All eleven analytes can be separated in a fused-silica capillary (63 cm to the detector, I.D. 75 microns) at 20 kV in a 0.02 mol 1(-1) phosphate-borate buffer (pH 8.0-9.0) with a separation factor > or = 1. The values of the Offord parameter calculated for individual nucleotides predict that monophosphates will migrate faster than triphosphates, and in turn triphosphates will precede diphosphates. By analogy, faster electroosmotic mobility (lower electromigration) of purine nucleotides (AP, GP) can be explained by a more voluminous structure of purine derivatives (two aromatic rings as compared to pyrimidines). Generally speaking, all compounds separated follow the Offord equation assuming that the triphosphate derivatives are ionized to the third degree forming HL(3-) anions. This assumption is in agreement with the current knowledge about protolytic equilibria of polyphosphates. The only exception to this rule is faster migration of guanosine-5'-triphosphate (GTP) preceding uridine-5'-monophosphate (UMP) which is ascribed in part to the larger molecule of GTP and the two additional OH-groups bound to the pyrimidine ring of UMP.

Enantiomer separation of dihydropyridine calcium antagonists with cyclodextrins as chiral selectors: structural correlation.

Native and substituted cyclodextrins (CDs) were used as chiral selectors both in high-performance liquid chromatography and capillary electromigration separations (HPCE and MEKC). Chromatographic data of five dihydropyridine calcium antagonists obtained on three beta-CD chiral stationary phases in reversed-phase mode were compared with those of capillary electrophoresis using beta-CDs in the presence and absence of sodium dodecyl sulfate (SDS). Competition of separated compounds with SDS molecules for penetration into the CD cavity can limit their necessary interaction with the chiral selector and consequently even preclude enantiomer separation. Some insight into this problem can be brought about by comparing the experimental data with computer-aided energy minimization of CD-solute and CD-SDS inclusion complexes.