On-line coupling of size exclusion and capillary zone electrophoresis via a reversed-phase C18 trapping column for the analysis of structurally related enkephalins in cerebrospinal fluid

On-line coupled analytical techniques can be advantageous in the assay of smaller peptides in complex biological matrices such as plasma, cerebrospinal fluid (CSF) and tissues. The present study shows the feasibility of a recently developed system, consisting of a size-exclusion chromatographic (SEC) separation followed by a trapping procedure on an RP18 microcolumn with subsequent elution of the trapped fraction and separation by capillary zone electrophoresis (CZE) for the quantification of structural-related peptides in biological matrices, as demonstrated for a number of enkephalins in CSF. After SEC separation of the enkephalins from large proteins present in CSF a heart-cut of 200 nuL, containing the enkephalin peak, is taken, concentrated on the RP18 microcolumn and, after elution of the enkephalins with 80% acetonitrile, a fraction of the eluate is electrokinetically injected into the CZE system, where stacking and separation is achieved. The degradation of the peptides, caused by endogenous peptidases in the matrix, is sufficiently inhibited with imipramine HCl. The assay has a satisfactory linearity and intraday (9.70-16.3%) precision considering the complexity of this multidimensional separation system. The sensitivity of the method, with a concentration limit of quantification of 2.5 nug/mL, is comparable with other CZE assays for peptides and sufficient for the quantification of peptide drugs in biological matrices.     

Determination of chloramphenicol and its monosuccinate ester in piglet plasma using HPLC

Summary A method for the determination of chloramphenicol and its monosuccinate ester in piglet plasma is described. It involves precipitation of plasma proteins by addition of methanol to the plasma sample, followed by injection of the supernate onto the column. Chloramphenicol and its ester are separated using a LiChrosorb RP-18 column and phosphate buffer pH 4.9 containing 30% methanol as eluent. Determination of chloramphenicol and its monosuccinate ester in pig plasma was made with a precision of 2.6% and 2.4% (relative standard deviation) at drug levels of 2 and 4 g·cm^–3, respectively. Stability data for the chloramphenicol monosuccinate ester are also presented.     

Separation of D- and L-DOPA as their L-leucine derivatives by reversed-phase HPLC

Summary The separation and quantitation of D-DOPA as an impurity in L-DOPA is described. The method involves formation of the diastereomeric derivatives of DOPA with tert-butoxycarbonyl-L-leucine hydroxysuccinimide ester. Separation of the diastereomers was performed by reversed-phase liquid chromatography on LiChrosorb RP-18 and acetonitrile-phosphate buffer with addition of N,N-dimethyloctylamine as the mobile phase. The reproducibility of the method was 5.9% (relative standard deviation) at D-DOPA levels of 0.5% in L-DOPA drug substance.     

Gas chromatographic determination of xanthinol in plasma

The determination of xanthinol in plasma is described. After extraction of the drug, together with the internal standard (papaverine hydrochloride), the extract is evaporated to dryness and the drug is derivatized with acetic anhydride for chromatography. The method is linear for 2–100μg ml^-1 ; the coefficient of variation is 3% and the recovery 80%. The resulting stable solution allows large numbers of samples to be processed with an automatic injector.     

Calcium adsorption and displacement: characterization of lipid monolayers and their interaction with membrane-active peptides/proteins

Background  

The first target of antimicrobial peptides (AMPs) is the bacterial membrane. In the case of Gram-negative bacteria this is the outer membrane (OM), the lipid composition of which is extremely asymmetric: Whereas the inner leaflet is composed of a phospholipid mixture, the outer leaflet is made up solely from lipopolysaccharides (LPSs). LPS, therefore, represents the first target of AMPs. The binding and intercalation of polycationic AMPs is driven by the number and position of negatively charged groups of the LPS. Also, proteins other than cationic AMPs can interact with LPS, e.g. leading eventually to a neutralization of the endotoxic effects of LPS. We compared different biophysical techniques to gain insight into the properties of the electrical surface potentials of lipid monolayers and aggregates composed of LPSs and various phospholipids and their interaction with peptides and proteins.     

Supersymmetric descendants of self-adjointly extended quantum mechanical Hamiltonians

We consider the descendants of self-adjointly extended Hamiltonians in supersymmetric quantum mechanics on a half-line, on an interval, and on a punctured line or interval. While there is a 4-parameter family of self-adjointly extended Hamiltonians on a punctured line, only a 3-parameter sub-family has supersymmetric descendants that are themselves self-adjoint. We also address the self-adjointness of an operator related to the supercharge, and point out that only a sub-class of its most general self-adjoint extensions is physical. Besides a general characterization of self-adjoint extensions and their supersymmetric descendants, we explicitly consider concrete examples, including a particle in a box with general boundary conditions, with and without an additional point interaction. We also discuss bulk-boundary resonances and their manifestation in the supersymmetric descendant.