Simultaneous analysis of disopyramide and quinidine in plasma by high-performance liquid chromatography

A new reversed-phase high-performance liquid chromatographic method allowing simultaneous measurement of plasma concentrations of disopyramide and quinidine is described. Disopyramide and quinidine were separated on a reversed-phase column using 0.05 M phosphate buffer (pH 3.0)—acetonitrile (73:27, v/v), as mobile phase and the peaks were monitored by UV absorbance at the wavelengths of 254 and 325 nm. The drugs were extracted from alkaline plasma with chloroform containing the internal standard. The organic phase was evaporated to dryness and the residue was redissolved in a small volume of the mobile phase before analysis by high-performance liquid chromatography. The method is convenient and reliable in routine monitoring of both drugs.     

Analysis of developmentally homogeneous neural crest cell populations in vitro: I. Formation,morphology and differentiative behavior

When early embryonic quail neural tubes are dissected free from surrounding tissues and placed in culture, small stellate neural crest cells usually migrate from the explant onto the substratum. This outgrowth has been reported to consist of a mixture of cells, some of which undergo melanogenesis, while the rest remain unpigmented. We have, in contrast to earlier observations, obtained a spatial separation of the two phenotypes. In these cultures the primary outgrowth of migrating cells remained almost free of pigment-forming cells, whereas small spherical clusters containing several hundred pigment-forming cells appeared on the explanted neural tubes. Whether the clusters remained with the tube explants or were subcultured, all cluster cells differentiated into melanocytes. Prior to melanogenesis, the appearance of the cultured cells from a cluster was indistinguishable from the cells in the outgrowth. The clusters provide a source of neural crest cells, that (1) can be easily obtained in comparatively large numbers, (2) is not contaminated with any other cell type, (3) can be isolated before the onset of differentiation, and (4) is developmentally homogeneous. Thus, the cluster population is well suited for many types of experiments, such as the identification of specific environmental factors that might control neural crest cell differentiation.     

Radioimmunoassay for the detection and quantitation of bruceantin

A radioimmunoassay for a new anticancer drug, bruceantin, has been developed using [³H]acetylbruceantin and antibody induced by immunizing rabbits with succinylbruceantin-bovine serum albumin conjugates. [³H]Acetylbruceantin was synthesized by reacting bruceantin with [³H]acetyl anhydride. The assay is simple and reproducible. The standard curve was linear on a logit-log plot, and the lower limit of sensitivity of the assay was 1 ng/ml. Using this assay, drug levels were easily determined in tissues of experimental animals following bruceantin administration. The assay procedure does not require sample extraction for plasma, urine, and bile. Bruceantin in other tissues can be extracted quantitatively with ethanol before being measured by the radioimmunoassay.     

Kälteresistenz der Fichte

Fichtenchloroplasten durchlaufen während eines Jahres saisonbedingte charakteristische Struktur- und Funktionswechsel: Frühjahrschloroplasten, die in den alten Nadeln kurz vor und während des Knospenaustriebes gefunden werden, sind sehr groß und so sehr mit Stärke erfüllt, daß man sie als Amyloplasten bezeichnen kann. Beim Aufbau der neuen Nadelgeneration wird diese Stärke verbraucht, und es entwickeln sich aus den Amyloplasten wieder die photosynthetisch aktiven Sommerpiastiden mit einem gut entwickelten Membransystem. Während der Frosthärtung und insbesondere während der Frostperiode treten die Chloroplasten in ihrer Winterform auf: amöboide, an einer Stelle der Zelle konzentrierte, stark aufgequollene Plastiden mit einem aufgelockerten und zum Teil reduzierten Thylakoidsystem. Der Strukturwandel der Chloroplasten wird — wie die Frostresistenz selbst — durch exogene Faktoren (Tageslänge, Temperatur) ausgelöst und kann auch durch artifiziell veränderte Umgebung zu unnatürlichen Zeitpunkten ausgelöst werden. Begleitet wird der Strukturwandel von einer Veränderung der CO₂-Fixierungsrate der Fichtennadeln, die ihrerseits auf Veränderungen der photochemischen Aktivität der Chloroplasten zurückzuführen ist. In Frostexperimenten konnte gezeigt werden, daß die Frosthärtung Reaktionen auf zwei verschiedenen Ebenen auslöst: 1. Produktion von kolligativ wirksamen Membranschutzstoffen sowie 2. einer Veränderung der chemischen Zusammensetzung der Chloroplastenmembranen; zumindest letztere führt offensichtlich zu einer Verringerung der Photosyntheseleistung. Frostschädigung der Chloroplasten tritt bei nicht entsprechend stark gehärteten Nadeln auf, jedoch nicht durch direkten Einfluß der tiefen Temperatur auf das Thylakoidsystem als vielmehr durch Freisetzung membranschädigender Substanzen durch Permeabilitätsverlust plasmatischer Membranen. Wir danken Herrn Prof. Dr. O. Kandler für anregende Diskussionen und für die kritische Durchsicht des Manuskriptes. Der Deutschen Forschungsgemeinschaft wird für die finanzielle Unterstützung dieser Arbeit gedankt.     

Improved basal medium for Y-1 mouse adrenal cortex tumor cells in culture

Summary An improved basal medium is presented that requires only minimal supplementation with dialyzed fetal bovine serum or bovine serum albumin and fetuin to be comparable to Ham's F-10, which requires 15% horse serum (HS) and 2.5% fetal bovine serum (FBS) for the growth and function of Y-1, mouse adrenal cortex tumor, cells. Cell monolayers maintained for up to 2 weeks without any protein supplementation have retained their steroid response to ACTH. The medium differs from Ham's F-10 in its buffer composition and higher calcium-ion concentration. This medium should be a useful adjunct to studies pertaining to steroid and lipid intermediary metabolism, the retention of a specialized physiological function in a chemically defined medium, and the mechanism of hormonal response. Supported by the Medical Research Service of the Veterans Administration.     

Biochemical and genetic analysis of a mutant with altered alkaline phosphatase activity in Dictyostelium discoideum

Alkaline phosphatase is one of several enzymes that accumulate in a temporally regulated sequence during the development of Dictyostelium discoideum. These enzymes can be used to monitor specific gene expression; moreover, isolation and analysis of mutations in the structural gene(s) can serve to indicate some of the essential steps in programmed synthesis and morphogenesis. A mutation (alpA) which affects the activity and substrate affinity of alkaline phosphatase was isolated in D discoideum using a procedure for screening large numbers of clones. Alkaline phosphatase activity at all stages of vegetative growth and development was altered by the mutation. Several physical properties of the enzyme from growing cells and developed cells were compared and found to be indistinguishable. It is likely that a single enzyme is responsible for the majority of alkaline phosphatase activity in growth and development. The mutation is coexpressed in diploids heterozygous for alpA and maps to linkage group III. One of the haploid segregants isolated from these diploids carries convenient markers on each of the six defined linkage groups and can be used for linkage analysis of other genetic loci.     

Growth of “Seeded” Cellulolytic Enrichment Cultures on Mesquite Wood

Eleven enrichment cultures were developed by a “seeded” enrichment culture technique, and one was developed by a simple enrichment technique. The seeded enrichment, the pure “seed,” and the simple enrichment cultures were compared during growth on mesquite wood, cotton, carboxymethylcellulose, and cellobiose. All of the enrichment cultures were cellulolytic and exceeded the pure seed cultures in mesquite wood hydrolysis and/or viable cell count. Yeast extract improved, but was not essential for, growth of the seeded enrichment cultures on carboxymethylcellulose. Two of the seeded enrichment cultures, CAD5 and CAD11, grew best at 37°C and pH 7.0 on mesquite wood. A 1.0% (wt/vol) wood concentration was optimum for their growth.     

Pyruvate metabolism after in vivo exposure to oral arsenic

This study investigated altered pyruvate metabolism after prolonged oral arsenic exposure. Male rats were given access to deionized drinking water containing 0, 40 or 85 ppm sodium arsenate (As⁵⁺) for 3 weeks. Respiration studies with mitochondria isolated from treated animals indicated decreased state 3 respiration (with ADP) and decreased respiratory control ratios (RCR) for pyruvate/malate-mediated respiration, but not for succinate-mediated respiration, as compared to control respiration values. In addition, pyruvate dehydrogenase activity was measured, in both liver and intestine, before and after Mg-activation in vitro. After 3 weeks, the effects of arsenic at the highest dose level were pronounced on the basal pyruvate dehydrogenase activity (before activation) as well as the total pyruvate dehydrogenase (after activation). The inhibition of pyruvate dehydrogenase activity both before and after Mg-activation suggests an arsenic effect on mitochondrial pyruvate metabolism which, in part, involves inhibition of pyruvate decarboxylase. Evidence is also presented which may indicate an arsenic effect on the kinase and/or phosphatase which regulate pyruvate dehydrogenase activity.