Biomedical subjects
Publications and source records attributed to C F Poole.
The principal methods for obtaining quantitative information from separations performed by planar chromatographic techniques are reviewed. Recent advances in obtaining structural information for sample identification of separated components are also discussed. Reasonable expectations concerning future developments in densitometry are made.
An evaluation of extraction procedures, liquid-liquid distribution systems, Sep-Pak cartridges, liquid-solid chromatography using silica, alumina and chemically modified silica packings (acid-base treated ethylammonium nitrate and picric acid impregnated), macroreticular resins and gel permeation columns for the analysis of polycyclic aromatic hydrocarbons (CPAHs) in environmental samples by thin-layer chromatography is discussed. For particulate samples solvent extraction using a Soxhlet apparatus or ultrasonication was found to be preferable to sublimation and liquid-liquid distribution between hexane and dimethyl sulfoxide followed by silica gel column chromatography was the preferred method for sample clean-up. Using this procedure enabled six PAHs (anthracene, fluoranthene, benz[a]anthracene, perylene, pyrene, and coronene) to be determined quantitatively in urban air particulate, diesel engine exhaust particulate, laboratory ventilator dust, household dust, river water, and tea samples. The PAHs were identified by coincidence of retention between the sample and standards in the same chromatographic system and by adequate agreement with standards for their normalized emission response ratios. The two-point calibration method was used for quantitation. Good agreement for the concentration of PAHs in the air particulate and diesel particulate extracts with published data using gas chromatography-mass spectrometry and high-performance liquid chromatography was found.
High performance thin-layer chromatography (HPTLC) with fluorescence scanning densitometry was used for the quantitative determination of polycyclic aromatic hydrocarbons in the soluble organic fraction of air particulate samples. A method using normalized emission response ratios was developed to determine sample identity and to test for peak homogeneity. To preserve the high sample throughput of HPTLC, the two-point calibration method was used for quantitation. The principal advantages of HPTLC as a screening technique for environmental samples are its low cost, methodological simplicity, high sample throughput, and the ability to analyze crude samples with a minimum amount of sample cleanup.
A comparison is made of different stationary phases (silica gel, cellulose, acetylated cellulose, and 3-aminopropyl-, ethyl-, octyl-, octadecyl-, and diphenylsilanized silica gel), mobile phases, and development techniques (conventional, continuous, and multiple) for the separation of environmentally important polycyclic aromatic hydrocarbons by high-performance thin-layer chromatography. Reversed-phase separations using octadecylsilanized silica gel and methanol-water or methanol-water-based ternary solvent systems were found to give the best separations. A new development technique, multiple development with application of the mobile phase further up the plate at each successive development, provided superior resolution of the polycyclic aromatic hydrocarbons compared with conventional multiple development.
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Acrylamide in biological samples can be determined by gas chromatography with electron-capture detection after conversion to its 2,3-dibromopropionamide derivative. The derivatization is carried out in aqueous solution, plasma or tissue homogenates by ionic bromination and the reaction conditions and sample clean-up are described. The detection limit corresponds to 9.5.10(-12)g of acrylamide on column or 8.4.10(-9)g in the final biological extract (0.5 ml). The overall recovery of acrylamide spiked samples at the nanogram level exceeds 80%. It was found that the accumulation of free acrylamide in the sciatic nerve distal region of rats intoxicated with acrylamide was less than 2-8 ppm.
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The organic volatile constituents of biological fluids contain clinically useful diagnostic information for the recognition of metabolic disorders in man. To gain access to this information, it was necessary to develop the methodology for reproducibly stripping the trace concentrations of volatiles from biological fluids (dynamic headspace, gas phase-stripping, solvent extraction, and the transevaporator technique), to separate the complex extracts by high-resolution capillary column gas chromatography, and to develop computer-aided data-handling and pattern-recognition techniques for analyzing the immense amount of information generated. The normal and pathological organic volatiles identified by gas chromatography--mass spectrometry in urine, serum, and breast milk are tabulated. Clinical applications of the above techniques to the study and diagnosis of diabetes mellitus, respiratory virus infection, renal insufficiency, and cancer are described.
Guaifenesin after extraction from plasma with an organic solvent can be selectively derivatized with 2,4-dichlorobenzeneboronic acid and determined by gas chromatography with electron-capture detection. The detection limit for guaifenesin was 15 ng ml-1 for a 2.0-ml plasma sample. The mass spectra of the boronate derivatives of guaifenesin and mephenesin, used as internal standard, show good molecular ions with characteristic modes of fragmentation useful for their identification.
The reaction between 2,4-dichlorobenzeneboronic acid, 3,5-bis(trifluoromethyl)benzeneboronic acid or the 1,3-propanediamine derivatives of the boronic acids and the 3-isopropylaminopropan-2-ol side chain of the beta-blocking drugs occurs essentially as an on-column thermal reaction in the gas phase. Derivatization of the side chain of the beta-blocking drugs by transboronation is shown to be the method of choice for general application as it affects the detector background signal very little compared to the use of the boronic acid itself. The transboronation reaction can be used for the determination of alprenolol in plasma extracts with a detection limit of 2.5 ng ml-1 using the electron-capture detector. The ultimate sensitivity of the method is limited by the detector background signal resulting from some column decomposition of the transboronation reagent.
The profiles of volatile metabolites in serum samples from normal individuals and from individuals with diabetes mellitus with varying degrees of polyneuropathy have been studied. The transevaporator procedure was used to obtain sample extracts which were chromatographed on a highly efficient glass column coated with Silar 10C (106 m x 0.25 mm I.D.). Differences in profiles between normal subjects and diabetic subjects on no drug therapy were noticed. However, correlations between the severity of the neuropathy and the concentrations of certain ketones could not be established. Compounds present both in diabetic and normal sera have been identified by mass spectrometry.
The response of the electron-capture detector to organic compounds is poorly defined, and the steroids are no exception to this observation. For those steroids which are naturally electron-capturing, the structures of the electrophores will be defined. Other steroids can be made electron-capturing by the formation of appropriate derivatives. Some new or infrequently used reagents for this purpose (flophemesyl ethers, t-bulflophemesyl ethers, pentafluorophenylhydrazone derivatives and halogen-substituted aromatic boronic acids) are described.
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Using a transevaporator sampling technique, the volatile profiles from 70 microliter of serum were obtained by capillary-column gas chromatography. The complex chromatograms were interpreted by a combination of manual and computer techniques and a two-peak ratio method devised for the classification of normal and virus-infected sera. Using the K-nearest neighbor approach 85.7% of the unknown samples were classified correctly. Some preliminary results indicate the possible use of the method for the assessment of virus susceptibility.
Selenium can be determined quantitatively in biological samples after nitric acid-magnesium nitrate digestion and formation of 5-nitropiazselenole, by extraction into toluene for gas-liquid chromatography with electron-capture detection. The method is suitable for the determination of selenium in orchard leaves, bovine liver and human placenta, hair, blood and urine.
A simple method has been devised for the analysis of di-(2-ethylhexyl)phthalate (DEHP) in human placenta which relies entirely on three rapid partition steps prior to gas chromatography with electron capture detection. The use of adsorption chromatography with all its attendant disadvantages for sample clean-up is eliminated in this procedure. Placental samples taken from women who had given birth to a normal baby contained 0.06 +/- 0.02 ppm of DEHP. The principal limitation of this method, and of any others, is the high blanks given by laboratory equipment and solvents. The reduction of contamination to workable levels is described.
The hydroxyl groups of 20-hydroxyecdysone react with trimethylsilylimidazole with varying ease, in the positional order 2,3,22,25 greater than 20 greater than 14. The 14alpha-hydroxyl group can only be silylated under forcing conditions. Confusion in silylation procedures has been caused by failure to recognize incomplete reaction. The conclusions are supported by mass spectra. In the presence of a catalyst, and absence of a 14alpha-oxy substituent, enol ethers are readily formed, but the rate is considerably reduced with a C-14 substituent present.
Cholest-7-en-6-one, 14alpha-hydroxycholest-7-en-6-one, 2beta, 3beta, 5alpha-trihydroxycholest-7-en-6-one and ten other related compounds have been synthesized by efficient routes from cholesterol. The named compounds possess some of the characteristics of the insect moulting hormones, ecdysones.