Determination of Poly-beta-Hydroxybutyrate and Poly-beta-Hydroxyvalerate in Activated Sludge by Gas-Liquid Chromatography

A convenient gas-liquid chromatography procedure to quantify poly-beta-hydroxybutyrate and poly-beta-hydroxyvalerate in activated sludge was developed by combining lyophilization of the samples, purification of the chloroform phase by water reextraction, and the use of capillary columns. With a flame ionization detector the sensitivity was estimated at 10 g/liter.     

Combination biotherapy utilizing interleukin-2 and alpha interferon in patients with advanced cancer: a National Biotherapy Study Group Trial.

The National Biotherapy Study Group (NBSG) conducted a broad phase II trial using interleukin-2 (IL-2) by continuous infusion and alpha interferon (IFN) subcutaneously in 267 patients with a variety of advanced cancers, including 29 with breast cancer, 89 with renal cancer, and 69 with melanoma. IL-2 [18 million international units (MIU)/m2] was given by continuous infusion for 108 hours with 3 mu/m2 subcutaneous IFN every other day during the IL-2 infusion. The patients were treated for 1 week followed by a 2-week rest. After two cycles of treatment, patients were evaluated for response. Of the 237 patients evaluable for response, 20 (8%) had a complete or partial response and 128 (54%) were stable. Therefore, 62% of the evaluable patients were nonprogressive during the first 90 days of IL-2/IFN therapy. The objective response rate was 11% in melanoma, 7% in renal cancer, 14% in breast cancer, and 3% in patients with a variety of malignancies for an overall response rate of 7% in these patients with advanced cancer. The patients were treated on a general medical ward and tolerated treatment well with fatigue and fever being nearly universal. Dyspnea, pruritus, chills, and elevated creatinines were frequent but less common. This combination biotherapy regimen has minimal activity in a variety of advanced cancers and must be compared with the best existing chemotherapy for each cancer type in randomized, prospective trials.     

Adhesion, penetration and intracellular replication of Legionella pneumophila: an in vitro model of pathogenesis

Legionella pneumophila attached to, penetrated and replicated within the three eukaryotic cell lines, MRC-5, HEp-2 and Vero. Multiplication occurred rapidly in these cells for an initial 48 h after inoculation and declined thereafter. Infected MRC-5 cell monolayers developed lytic-type cytopathic changes, with organisms being readily released. HEp-2 cells showed a more chronic infection, with slowly developing granular changes in the monolayers, and slow release of intracellular bacteria. In Vero cells, organisms were released rapidly along with a more progressively developing granular cytopathic effect in the monolayers. L. pneumophila was unable to grow in cell-free culture fluids. Uptake and intracellular development was similar for each cell type, and was initiated by 'bacteriopexis', a process in which the organisms bound via receptors and were surrounded by cellular microvilli which eventually fused, leading to bacterial engulfment. Replication of organisms in vacuoles within the cytoplasm of infected cells was confirmed by thorium labelling. These vacuoles were lined with ribosomes and, at the early stages of intracellular development, were found in close proximity to mitochondria, cytoplasmic filaments and banded enclosures. Ruthenium red staining showed that acid mucopolysaccharide capsular material was not present on these organisms during the attachment process or intracellular phase. Organism release was by lysis of the infected cells.     

Anaerobic acidogenesis of primary sludge: the role of solids retention time

This research investigates the effect of solids retention time (SRT) on the acid-phase anaerobic digestion of primary sludge. A series of experiments were conducted using two continuous-flow 3-L units with the following configuration: a completely mixed reactor (CMR) with clarifier and solids recycle and an upflow anaerobic sludge blanket (UASB) reactor. Results show that C(2) to C(5) volatile fatty acids (VFA) were the predominant compounds formed. At a constant hydraulic retention time (HRT) of 12 h, variation in SRT from 10 to 20 days resulted in a slight increase in VFA production in both systems, but at a shorter SRT (5 days) a drastic drop in acid production was observed. In addition, the percent distribution of VFA was to some extent affected by the change in SRT. On the other hand, organic matter degradation [measured by the chemical oxygen demand (COD) specific solubilization rate or the percent volatile suspended solids (VSS) reduction] appeared to be independent of SRT, at least in the range investigated. The percent soluble COD in the form of VFA, however, increased steadily with increasing SRT, approaching the 90% level at 20 days. The remaining soluble COD in the effluent from these systems may be mainly attributed to metabolic intermediates and unused soluble substrate. (c) 1994 John Wiley & Sons, Inc.     

Isolation and characterization of plasma membrane-associated cortical granules from sea urchin eggs

Cortical granules, which are specialized secretory organelles found in ova of many organisms, have been isolated from the eggs of the sea urchins Arbacia punctulata and Strongylocentrtus pupuratus by a simple, rapid procedure. Electron micropscope examination of cortical granules prepared by this procedure reveals that they are tightly attached to large segments of the plasma membrane and its associated vitelline layer. Further evidence that he cortical granules were associated with these cell surface layers was obtained by (125)I-labeling techniques. The cortical granule preparations were found to be rich in proteoesterase, which was purified 32-fold over that detected in a crude homogenate. Similarly, the specific radioactivity of a (125)I-labeled, surface glycoprotein was increased 40-fold. These facts, coupled with electron microscope observations, indicate the isolation procedure yields a preparation in which both the cortical granules and the plasma membrane-vitelline layer are purified to the same extent. Gel electrophoresis of the membrane-associated cortical granule preparation reveals the presence of at least eight polypeptides. The major polypeptide, which is a glycotprotein of apparent mol wt of 100,000, contains most of the radioactivity introduced by (125)I-labeling of the intact eggs. Lysis of the cortical granules is observed under hypotonic conditions, or under isotonic conditions if Ca(2+) ion is present. When lysis is under isotonic conditions is induced by addition of Ca(2+) ion, the electron-dense contents of the granules remain insoluble. In contrast, hypotonic lysis results in release of the contents of the granule in a soluble form. However, in both cases the (125)I-labeled glycoprotein remains insoluble, presumably because it is a component of either the plasma membrane or the vitelline layer. All these findings indicate that, using this purified preparation, it should be possible to carry out in vitro studies to better define some of the initial, surface-related events observed in vivo upon fertilization.     

Comparative induction of unscheduled DNA synthesis by physical and chemical agents in non-proliferating primary cultures of rat hepatocytes

The incorporation of [³H]thymidine into DNA due to unscheduled DNA synthesis (UDS) induced by N-OH-2-acetylaminofluorene (N-OH-AAF), aflatoxin B₁ (AFB₁), ethyl methanesulfonate (EMS) and ultra-violet light was quantitated by autoradiography and by scintillation spectrometry on acid precipitable macromolecules or DNA insolated by isopycnic banding in cesium chloride (CsCl). Dose-dependent increases in UDS due to N-OH-AAF and AFB₁ treatment were found. Only 2-fold increases at the highest dose levels were found, however, when incorporated [³H]thymidine was quantitated by scintillation spectrometry. Seven, 11, and 25-fold increases in UDS induced by AFB₁, N-OH-AAF and ultra-violet light, respectively, were found when incorporated [³H]thymidine was quantitated by autoradiography, indicating a high sensitivity for detecting ‘long patch’ repair by this technique. Scintillation spectrometry was completely ineffective in detecting EMS-induced UDS, whereas autoradiography demonstrated a small, but significant induction in [³H]thymidine incorporation at high dose levels. The non-proliferative nature of the primary hepatocyte prohibits the uniform radioactive prelabeling of DNA, necessary in other techniques, for the detection of ‘short patch’ repair induced by compounds such as EMS. Therefore, the sensitivity of the primary cultured rat hepatocyte in conjunction with UDS for detecting DNA damage caused by mutagens and carcinogens which induce ‘short patch’ repair may be limited to the autoradiographic analysis of the unscheduled incorporation of [³H]thymidine.     

Human natural cell-mediated cytotoxicity

Summary The contribution of natural cell-mediated cytotoxicity (NCMC) to tumor-directed cytotoxicity is unknown. This study was undertaken to correlate changes in NCMC to a single target cell (K-562) with responses to therapy in patients with small cell carcinoma of the lung, resectable stage I non-small cell carcinoma of the lung, and stage I and II melanomas. Data from these studies suggested that these patients have depressed levels of NCMC to K-562 compared with a normal age-matched control population. However, NCMC levels appear to fluctuate with tumor burden, being highest in patients with large tumor masses and lowest in patients with no clinical evidence of tumor following a successful response to therapy.     

Comparison of the bile salts of frogs with those of their tadpoles. Bile-salt changes during the metamorphosis of Rana Catesbeiana Shaw.

1. The bile salts of three frog species of the genus Ptychadena and of Rana catesbeiana have been compared with those of their tadpoles. For R. catesbeiana comparison was made of the bile salts in at least ten of the recognized stages of tadpole metamorphosis. 2. In all cases, adult bile salts were more complex than those of the tadpoles. 3. In R. catesbeiana after stage 18, 26-deoxy-5 alpha-ranol was hydroxylated to form 5 alpha-ranol (27-nor-5 alpha-cholestane-3 alpha, 7 alpha, 12 alpha, 24 xi, 26-pentol) and at least two other bile alcohols appeared in solvolysed bile salts. 4. Tadpole bile salts were not found to be biochemically more primitive than those of fully metamorphosed frogs; in some, but not all, cases tadpole bile alcohols could be regarded as biochemical precursors of those in the adult frogs. 5. Detailed evidence for the structure of the bile salts from mass-spectral fragmentation patterns has been deposited as Supplementary Publication SUP 50097 (2 pages) at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1978) 169, 5.