Robert W. Walker

Coprostanol as an indicator of fecal pollution

Coprostanol is a fecal sterol formed by microbial reduction of cholesterol in the intestines of man and higher animals. The fact that this sterol is ubiquitously present in the feces of man and is excreted at the rate of about 1 g/capita/day has prompted investigators to suggest the use of this sterol as an indicator of fecal pollution. It has been found to be particularly useful in problem situations such as toxic or heated effluents and seawater environments where the use of the usual indicator bacteria is suspect. This review discusses the advantages and disadvantages of using coprostanol as an indicator sterol, details the results of numerous field studies, and examines the future prospects for this pollution indicator.

Toxicity and binding of copper, zinc, and cadmium by the blue-green alga, Chroococcus paris

The toxic effects and accumulation of the heavy metals, Cd, Cu, and Zn by the sheath forming blue-green alga Chroococcus paris were investigated. All three of the metals were bound rapidly. Approximately 90% of the total amount of the added metal was bound within 1 min. Further significant binding occurred at a slower rate. The maximum metal binding capacity, as determined by filtration studies, was determined to be 53, 120, and 65 mg g−1 dry algal weight for Cd, Cu, and Zn, respectively. Binding curves for the metals followed the Langmuir adsorption isotherm model. The amount of metal bound increased with increasing pH. Metal binding increased significantly when pH was increased from 4 to 7. Nearly all of the metal was found to be rapidly EDTA extractable. Metals were found to be increasingly toxic to growing cultures in the order, Zn, Cd, and Cu. All of the metals studied exhibited toxic effects at concentrations greater than 1.0 mg L−1. The lowest concentrations used which showed detectable toxicity were 0.1 mg L−1 for Cu and >0.4 mg L−1 for Cd and Zn.

The positional distribution of fatty acids in the phospholipids and triglycerides ofMycobacterium smegmatis andM. bovis BCG

Position 1 of the phospholipid and triglyceride fractions isolated fromMycobacterium smegmatis andM. bovis BCG was esterified principally with C18 related fatty acids (18∶0, 18∶1 and 19Br). Position 2 was occupied principally by C16 fatty acids. The third position of the triglycerides was esterified with a preponderance of C20+fatty acids. Seventysix per cent of position 3 fatty acids in BCG and 43% inM. smegmatis triglycerides contained fatty acids of greater than 20 carbon atoms.

Unsaturated fatty acids of mycobacteria

The double bond locations have been determined for the mono-unsaturated fatty acids, C14 to C26 ofM. smegmatis andM. bovis BCG. The 14∶1 and 16∶1 fatty acids fromM. smegmatis are principally Δ10, while the 17∶1, 18∶1 and 19∶1 fatty acids from both organisms are Δ9. In the case ofM. smegmatis, the 20∶1, 22∶1 and 24∶1 fatty acids are principally Δ11, Δ13 and Δ15, respectively, while the 22∶1, 24∶1 and 26∶1 fatty acids of BCG are principally Δ13, Δ15 and Δ17, respectively.

Cis-11-hexadecenoic acid fromcytophaga hutchinsonii lipids

The principal fatty acid (90+%) of the monoene fraction ofCytophaga hutchinsonii has been identified ascis-11-hexadecenoic acid. This fatty acid, which constitutes 29% of the total lipid fatty acids, is located principally in the phosphatidyl ethanolamine fraction.

The use of xad-2 resin for the analysis of coprostanol in water

Abstract The use of Amberlite XAD-2 resin column for extracting 5β-cholestan-3β-ol (coprostanol), a fecal sterol, from water was studied. It was found that the efficiency of the column extraction was comparable to that of the conventional liquid—liquid partitioning process. Maximum recovery from water samples depended upon pH of the sample, flow rate, resin mesh size, and concentration of coprostanol. Our data suggest that the use of Amberlite XAD-2 resin columns for the quantitation of coprostanol may be employed effectively to supplement the enumeration of coliform bacteria as an indicator of fecal pollution of water.

An XAD-1 column method for the rapid extraction of phytoplankton chlorophylls

Abstract A simple column method for the rapid filtration and extraction of phytoplanktonic chlorophylls has been developed. The method, tested on unialgal cultures (Chlorella vulgaris, Oocystis marssonii, Chlorococcum hypnosporum, Chlamydomonas moewusii, Oscillatoria tenius), hypereutrophic, mesotrophic and clean water field samples was found equivalent to or, in most cases, better than the conventional filtration/acetone extraction procedures. In the new procedure, samples were filtered through an XAD-1 resin (60–120 mesh) column. Chlorophylls were extracted from the algal cells with basic methanol which deposited these compounds in the top layer of the column. The chlorophylls were then eluted by benzene and quantitated for by spectrophotometry. One and one half hours was required for the complete analysis of a 1-l sample.

A solvent partitioning procedure for the separation of chlorophylls from their degradation products and carotenoid pigments

A simple liquid/liquid partitioning procedure was developed which employed aqueous acetonitrile and hexane, for the isolation of chlorophyll and pheophytin. This procedure separated these pigments from other interfering pigmented compounds. The efficacy of this solvent separation method was evaluated using commercially available chlorophylla, b, pheophytina, b, carotenoids, and algal pigment extracts. The recovery efficiencies of this solvent partitioning process for chlorophyll a and pheophytina have been shown to be 95–98% and 93–96%, respectively, furthermore, the chlorophylla fraction was practically free of any contaminating pigments. It appears that a more accurate assessment of chlorophylla and pheophytina can be accomplished employing liquid/liquid partitioning than with the present standard method.

Neutral lipid biosynthesis in Mycobacterium smegmatis

The biosynthesis of neutral lipids in Mycobacterium smegmatis was studied using cell free extracts. Maximum neutral lipid production was obtained when the reaction mixture (400 microliter) consisted of 0.25 M potassium phosphate buffer (pH 7.5), 0.125 mM oleoyl-CoA, 3.75 mM sn-glycerol-3-P, 10 mM MgCl2 and 1.85 mg bovine serum albumin. No magnesium dependency for the acylation of sn-glycerol-3-P was observed. A slight stabilizing effect seemed to occur due to this ion. The enzyme phosphatidate phosphohydrolase, on the other hand, was shown to be magnesium dependent. The activity of this enzyme also appeared to be stimulated by high concentration (0.75 to 1.25 mM) of ATP which enhanced lipid formation at all concentrations tested (0.25 to 3.75 mM). A heat-stable protective factor having a molecular weight less than 16 000 which caused a stimulatory effect on sn-glycerol 3-phosphate acyltransferase activity was found in the cell-free extracts. Preliminary experiments suggest that the factor might be polysaccharide in nature.

A simplified method for the simultaneous extraction of phytoplanktonic chlorophyll and fecal sterol from water

A simple column method employing Amberlite XAD-1 (60–120 mesh) resin for the simultaneous extraction of coprostanol and algal chlorophyll a was proposed and evaluated. The results showed that the efficiencies of the column extraction for these two water quality indicators were comparable to or better than the conventional hexane liquid/liquid partitioning and aqueous acetone extraction for the respective compounds. Furthermore, the column technique was time-saving and more economical.