A simple method for the quantitative determination of muramic acid

A simple method for microdetermination of muramic acid is elaborated. The method is based on the degradation of muramic acid to lactic acid, followed by degradation of the latter to acetaldehyde which can be determined colorimetrically with p-hydroxydiphenyl (PHD). A linear relationship exists between the concentration of muramic acid (up to 20 μg), and absorbance at 560 nm. Substances usually present in the hydrolysates of bacterial cell wall peptidoglycan do not interfere in the determination.     

A simple and inexpensive multiple tube system for thein situ measurement of phytoplankton production by the14C method

A cheap and simple multiple tube incubation system is described for the in situ measurement of phytoplankton production by the ¹⁴C method. Incubation takes place in Pyrex glass tubes sealed with rubber suba-seal caps which are suitable for field, laboratory and incubator work. Comparison of production in the tubes with that in standard pyrex bottles showed no significant difference during both field and laboratory incubations. The in situ suspension system is transported in units and can be assembled into ladders in the field.     

A simple method for production of arachidonic acid byMortierella alpina

Summary A filamentous fungus,Mortierella alpina was incubated aerobically on a medium made from potato paste and dextrose at 20 °C for 20 days. The yield of arachidonic acid reached 11.8 g/kg medium, which is more than 10 times higher than that by any other method reported previously. The highest content of the acid was 67.4% of the total fatty acids.     

Elongation of (omega-14C)oleic acid and (omega-14C)nervonic acid.

During feeding experiments with [omega-14C]oleic acid and [omega-14c]nervonic acid to adult rats, 14C-labelled C26, C28 and C30 fatty acids were recovered from the intestinal mucosa, liver, plasma, kidney and stools. The structures of these fatty acids were determined by g.l.c., radio-g.l.c. and mass spectrometry. The Schmidt and Ginger degradation methods indicated that most of the 14C found in these extra-long fatty acids remained in the omega position. These radioactive extra-long fatty acids were found mainly in the polar lipids of rats killed 3 or 15 h after being fed on labelled oleic acid or nervonic acid. Rats killed 63 h later yielded only traces of these extra-long fatty acids. When the rats were given antibiotics or received the same radioactive fatty acids by intravenous injection, the labelled extra-long fatty acids could not be detected in any of the tissues. We conclude that they were probably synthesized by elongation of oleic acid and nervonic acid by intestinal micro-organisms (probably yeasts) and then absorbed by the intestinal mucosa.     

A differential staining method for mast cells and eosinophils in murine intestine,liver and spleen

The detection of eosinophils and mast cells in tissues has been problematic. In this article, Parviz Kermanizadeh, Paul Hagan and David Crompton describe a new differential staining technique for mast cells and eosinophils in murine tissues and compare it with the standard methods.     

A simple binding assay for the determination of low-density lipoprotein receptors in cell homogenates

A convenient binding assay has been developed for the determination of low-density lipoprotein (LDL) receptors in homogenates of cultured and freshly-isolated normal and malignant human cells. Cell homogenates were incubated with 125I-labeled LDL and the ligand bound to the homogenate particulates was separated from the unbound ligand by filtration. When the particulates of the homogenates were subsequently incubated with heparin, a fraction of the bound 125I-LDL was released. Previous studies on intact cells have shown that heparin exclusively releases LDL bound to its cell surface receptor. The heparin-sensitive binding of 125I-LDL to cell homogenate particulates represents LDL bound to its cell surface receptor as judged from the following criteria: (a) it was quantitatively similar to the heparin-sensitive binding of 125I-LDL to intact cells, (b) it showed a direct correlation to the receptor-mediated degradation of 125I-LDL by intact cells, (c) no heparin-sensitive binding could be detected in homogenates prepared from normal erythrocytes or from cultured fibroblasts from a patient with homozygous familial hypercholesterolemia (two types of cell lacking LDL receptors), (d) it was dependent on calcium and inhibited by EDTA, (e) it was susceptible to treatment with pronase, and (f) it was heat-labile. The assay developed should be of value in determining the number of LDL receptors in tissues, since it is far less time-consuming and requires less material than currently available methods.     

A simple method for measuring immunoglobulin synthesis in vivo in the spleen of chicks (Gallus domesticus)

1. Spleen immunoglobulin (IgG) synthesis was measured in vivo in chicks at 3 weeks of age by a large dose injection of labelled phenylalanine in combination with the isolation of IgG by immunological precipitation against anti-chicken IgG. 2. No appreciable amount of radioactivity was detected in serum IgG for the first 60 min of the isotope injection via the wing vein, indicating a minimum time lag necessary for the secretion of newly synthesized IgG into the circulation. 3. Synthesis rates of total spleen protein and spleen IgG were found to be 17.5 and 4.8 mg/day, respectively, suggesting that IgG synthesis would contribute to 27% of the total protein synthesis in the spleen of young chicks.     

A new method for the determination of 14C radioactivity in ketone bodies.

A reaction vessel is described consisting of a simple reaction chamber fixed to an ordinary liquid scintillation-counting vial which allows the direct determination of radioactivity in acetone as well as separately in the carboxyl and acetone moieties of acetoacetate and 3-hydroxybutyrate. Radioactivity and its distribution between carbon 1 and carbons 2 through 4 can be determined in total ketone bodies as well as sequentially in acetone, acetoacetate, and 3-hydroxybutyrate. Recoveries with labeled ketone bodies under a variety of analytical conditions are presented.     

A sensitive method for the separation and quantitation of (14C)proline and (14C)hydroxyproline from the collagen of rat uterus

A specific and sensitive method is described for the isolation and quantitation of [¹⁴C]proline and [¹⁴C]hydroxyproline from uterine collagen of the immature rat. Selectivity is achieved in this isolation by using a protease-free bacterial collagenase. There is complete release of hydroxyproline from uterine protein if the latter is suspended by sonication prior to treatment with collagenase. There is a consistent recovery of [¹⁴C]proline and [¹⁴C]hydroxyproline when they are added to protein hydrolysates of uterus and then subjected to the procedures required for their isolation and quantitation. It is possible using this method to determine the incorporation of [¹⁴C]proline into collagen of the rat uterus and to quantitate its conversion to [¹⁴C]hydroxyproline. Coupled with the colorimetric methods for proline and hydroxyproline, it is also possible to determine their specific activity.