The use of hydrocarbon analyses for environmental assessment and remediation

Battelle Ocean Sciences has developed an analytical approach to identify and’ quantify petroleum products, coal products, and individual hydrocarbon components at trace levels in complex environmental matrices. The hydrocarbon analysis strategy uses capillary gas chromatography/flame ionization detection for alkane and total oil analysis, combined with gas chromatography/mass spectrometry for polynuclear aromatic hydrocarbon analysis. The method provides environmentally realistic analyte detection limits (parts per trillion in water, parts per billion in sediments) and an analyte list that is designed specifically for petroleum and coal‐based products. Results are compared to a detailed computerized library of total, water‐soluble, and degraded hydrocarbon products. The systematic data interpretation strategy maximizes the accuracy of petroleum and coal product identification in environmental matrices and represents a vast improvement over standard EPA methodology.     

海洋石油降解微生物及其降解机理

微生物修复作为一种新型环保的生物修复技术,已成为海洋石油污染生物修复的核心技术。对海洋石油降解微生物的种类即细菌、蓝藻、真菌以及藻类进行了总结,对微生物对石油烃的降解途径与降解机理进行了综述。微生物降解烷烃的过程包括末端氧化、烷基氢过氧化物以及环己烷降解3种形式。微生物对芳香烃的降解是通过芳香烃被氧化酶氧化导致苯环开环来实现的。微生物对多环芳烃的降解是在单加氧酶或双加氧酶的催化作用下被最终降解为二氧化碳和水而被分解。并对影响石油烃降解微生物的因素包括温度、营养物质等因素进行了分析。     

Determination of polycyclic aromatic hydrocarbons in smoked meat products and smoke flavouring food additives

Formation, factors affecting concentrations, legal limits and occurrence of polycyclic aromatic hydrocarbons in smoked meat products and smoke flavour additives are briefly reviewed. The most used techniques such as thin-layer chromatography (TLC), gas chromatography (GC) and high-performance liquid chromatography (HPLC) are evaluated. Also, sample preparation, pre-separation procedures, separation and detection systems being used for determination are discussed with emphasis to latest development in applied food analysis and the chosen data regarding the concentration of polycyclic aromatic hydrocarbons in smoked meat products and smoke flavour additives are summarised.     

Characterization of bio-oil from induction-heating pyrolysis of food-processing sewage sludges using chromatographic analysis

In this study, gas chromatography–mass spectrometry (GC–MS) was used to analyze the pyrolytic bio-oils and gas fractions derived from the pyrolysis of industrial sewage sludges using induction-heating technique. The liquid products were obtained from the cryogenic condensation of the devolatilization fraction in a nitrogen atmosphere using a heating rate of 300 °C/min ranging from 25 to 500 °C. The analytical results showed that the pyrolysis bio-oils were very complex mixtures of organic compounds and contained a lot of nitrogenated and/or oxygenated compounds such as aliphatic hydrocarbons, phenols, pyridines, pyrroles, amines, ketones, and so on. These organic hydrocarbons containing nitrogen and/or oxygen should originate from the protein and nucleic acid textures of the microbial organisms present in the sewage sludge. The non-condensable devolatilization fractions were also composed of nitrogenated and oxygenated compounds, but contained small fractions of phenols, 1H-indoles, and fatty carboxylic acids. On the other hand, the compositions in the non-condensable gas products were principally carbon dioxide, carbon monoxide and methane analyzed by gas chromatography–thermal conductivity detector (GC–TCD).     

Profiles of polycyclic aromatic hydrocarbon metabolites after treatment with various inducers of microsomal rat liver monoxygenases (author's transl)

The microsomal oxidation of 12 frequently occurring environmental polycyclic aromatic hydrocarbons after incubation with rat-liver microsomes has been studied and their metabolites characterized by means of gas-liquid chromatography/mass spectrometry. The method enables the detection and characterisation of phenols, diols, triols, and tetrols as trimethylsilyl ethers beside the original hydrocarbons. Moreover, the induction properties of some carcinogenic and non-carcinogenic hydrocarbons (benz[a]anthracene, pyrene, chrysene, benzo[a]-pyrene, benzo[e]pyrene, benzo[b]fluoranthene, benzo[j]fluoranthene, benzo[k]fluoranthene) have been studied. Except pyrene and benzo[e]pyrene, all compounds investigated significant but different induction factors. The relevance of the induction for an estimation of the biological effect of environmental polycyclic aromatic hydrocarbons is discussed.     

Standard reference materials for chemical and biological studies of complex environmental samples.

Standard Reference Materials (SRMs) from the National Institute of Standards and Technology (NIST) are often used in methods development and interlaboratory comparison studies since they are homogeneous and readily available to the scientific community. SRM 1649 (urban dust/organics), SRM 1650 (diesel particulate matter), and SRM 1597 (complex mixture of polycyclic aromatic hydrocarbons from coal tar) are three environmental samples which have been used by the scientific community for these purposes. These SRMs were originally developed to assist laboratories in validating analytical procedures for the determination of polycyclic organic compounds in complex mixtures. In addition, these SRMs have been valuable for the comparison of methodologies for bacterial bioassays and the development of bioassay-directed fractionation and bioassay-directed chemical analysis techniques. Most recently these SRMs were chosen for use as test samples in a collaborative study coordinated by the World Health Organization--International Programme on Chemical Safety. This paper provides a summary of much of the work to date (published and unpublished) on the chemical and biological characterization of these three SRMs. Information regarding the availability of other NIST SRMs that might be useful for these types of studies are provided also.     

Review on the analysis of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine and its phase I and phase II metabolites in biological matrices, foodstuff and beverages

The heterocyclic aromatic amine, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), has been shown to be carcinogenic in rodents, mice and rats. Following phase I N-hydroxylation and phase II esterification PhIP exerts its carcinogenic effect by binding to DNA purines. Quantitative and qualitative analysis of its bioactivated metabolites as well as it detoxification products is important in studying its biological effects and inter- and intra-individual exposures. A review is presented with an extensive coverage of publications specifically reporting on the analysis of PhIP and its phase I and II metabolites in biological matrices, foodstuff and beverages. Analytical techniques such as liquid and gas chromatography coupled with various detection techniques (mass spectrometry, ultraviolet or fluorescence detection) were mostly applied. We conclude that since the initial identification of PhIP in 1986 a large set of assays has been developed for the analysis of PhIP and its phase I and phase II metabolites in a wide range of matrices, these included food products and biological samples such as plasma, urine and faeces. In addition, it was shown that numerous metabolites were recovered and identified. Thus, we conclude that liquid chromatography coupled to mass spectrometry is clearly the method of choice for sensitive qualitative as well as quantitative analysis with high selectivity and reaching lower quantification levels in the sub pg/mL range. The main aim of this review is that it can be used by other researchers as a resource for method development and optimization of analytical methods of PhIP and its carcinogenic or detoxification products.     

Determination of mixtures of benzo[a]pyrene, 2,6-dimethylnaphthalene and their metabolites by high-performance liquid chromatography with fluorescence detection

Nonradiometric techniques were used to identify and quantitate two aromatic hydrocarbons and several of their metabolites in biological samples. High-performance liquid chromatographic separation combined with ultraviolet fluorescence detection provided a sensitive and selective method of trace analysis. Metabolites of benzo[a]pyrene and 2,6-dimethylnaphthalene were prepared by in vitro incubation of these substrates, singly and together, with liver microsomes of coho salmon (Oncorhynchus kisutch). Individual metabolites were separated by high-performance liquid chromatography and quantitated at fluorescence excitation and emission wavelengths characteristic of the corresponding parent hydrocarbon. Future refinement of these techniques may allow the determination of more complex mixtures of xenobiotics and their metabolites in marine organisms.     

Specific assay for radiolabelled digoxin and its known apolar metabolites in biological fluids. I.

A specific assay method for radiolabelled digoxin and its known apolar metabolites in plasma, urine and saliva was developed. The assay permits the delineation of the pharmacokinetics of digoxin and its metabolites after single-dose administration of the drug to humans. Column chromatographic and solvent extraction procedures were used for the separation of apolar and polar compounds. Thin-layer chromatography was applied for the individual and specific assessment of digoxin and its apolar metabolites. Apolar and polar standards were used for quantitative assessments of all the procedures used. Accuracy and precision of the assay developed were evaluated in plasma, urine and saliva using biological samples spiked with known amounts of standards and by measuring replicates of biological samples obtained from pharmacokinetic studies with digoxin administration to humans.     

Chemical Composition of Essential Oils and Aromatic Waters from Different Italian Anthemis maritima Populations

The chemical composition of the essential oils and aromatic waters isolated from six Italian Anthemis maritima populations was determined by GC‐FID and GC/MS analyses. In total, 122 and 100 chemical compounds were identified in the essential oils and the aromatic waters, respectively. The main compound classes represented in the oils were monoterpene hydrocarbons, oxygenated monoterpenes, sesquiterpene hydrocarbons, oxygenated sesquiterpenes, and terpene esters. Multivariate chemometric techniques such as cluster analysis (CA) and principal coordinate analysis (PCO) were used to classify the samples according to the geographical origin. Statistical analysis allowed the attribution of the analyzed populations to different chemotype groups.     

The formation and occurrence of polynuclear aromatic hydrocarbons associated with food

Polynuclear aromatic hydrocarbons are common contaminants of processed food, usually at trace levels. These hydrocarbons are products of combustion and pyrolysis, and are present in petroleum and coal, and in products derived from them. Most polynuclear aromatic hydrocarbons are not carcinogenic, but some of them are, and a few are potent inducers of skin and lung tumors in mice. Their carcinogenic properties have not been fully explored, but they seem to be less potent by ingestion or inhalation, and they are known as a group to produce cancer in humans. The most effective carcinogens among them are those with 5 or 6 fused rings, and these tend to be less prevalent in mixtures than the 3- and 4-ring hydrocarbons, most of which are not carcinogenic. Sophisticated analytical methods, using solvent extraction and chromatography have been developed to detect and measure polynuclear aromatic hydrocarbons at levels of 1 in 10(9) (1 part per billion) or less, and these have been applied to the measurement of individual compounds in foods, as well as in products of combustion and pyrolysis. Wood smoke and smoked foods contain the carcinogenic benzo[a]pyrene at levels of 1 ppb, and other hydrocarbons; liquid smoke has lower levels. Crude vegetable oils have higher concentrations, but purified 'deodorized' oils have benzo[a]pyrene levels near 1 ppb. Sausages cooked over burning logs had as much as 200 ppb benzo[a]pyrene. Charcoal-broiled steaks and ground meat had benzo[a]pyrene concentrations up to 50 micrograms/kg, while less fatty pork and chicken had lower concentrations (up to 10 micrograms/kg). It was probable that the rendered fat dripped on to the hot charcoal and pyrolyzed to form quantities of polynuclear aromatic hydrocarbons, which rose with the smoke to deposit on the meat. Therefore, oven cooking or cooking with a heat source above the meat, or segregation of the meat from the smoke resulted in food containing negligible amounts of polynuclear aromatic hydrocarbons. Modifications of cookings practices accordingly would greatly reduce exposure to this group of carcinogens.     

Microbial petroleum degradation: application of computerized mass spectrometry.

An analytical procedure is presented for obtaining detailed characterization of petroleum hydrocarbons which undergo microbial degradation. The procedure includes column chromatographic separation and characterization of the resulting fractions by mass spectrometry and gas chromatography. The use of computerized low-resolution mass spectrometry is offered as a method for assessing microbial degradation of petroleum. This method provides information which cannot, at the present time, be obtained by other available analytical methods. Use of this method to evaluate degradation of a South Louisiana crude oil by a mixed culture of estuarine bacteria revealed that asphaltenes and resins increased by 28% after degradation, while saturates and aromatics decreased by 83.4% and 70.5%, respectively. Most of the normal and branched-chain alkanes were degraded (96.4%), but an increase in long-chain alkanes (C28-C32) after degradation was observed by gas-liquid chromatography. Susceptibility of cycloalkanes to degradation was less as the structure varied, i.e., 6-ring greater than 1-ring greater than 2-ring greater than 3-ring greater than 5-ring greater than 4-ring. Susceptibility of aromatic components to degradation decreased with increase in the number of rings, viz., monoaromatics greater than diaromatics greater than triaromatics greater than tetraaromatics greater than pentaaromatics. Aromatic nuclei containing sulfur were twice as refractory as non-sulfur analogs.     

Analysis of Diesel Fuel Contamination in Soils by Near-Infrared Reflectance Spectrometry and Solid Phase Microextraction-Gas Chromatography

A feasibility study was undertaken to determine whether the rapid, nondestructive analytical technology, near-infrared reflectance spectrometry (NIRS) could be used to predict total petroleum hydrocarbon (TPH) in contaminated soil. Hydrocarbon concentrations were determined on samples of diesel-contaminated soils by the solid phase microextraction-gas chromatography (SPME-GC) method. The same samples were then scanned for near-infrared reflectance spectrometry over the wavelength range 1100 to 2498 nm. Calibrations were developed between the NIR spectral data and the reference SPME-GC chemical data using stepwise multiple linear regression. Linear regression relationships between NIR-predicted TPH concentrations and reference data had r² of 0.68 and 0.72. These results indicate that the combination of NIRS and SPME-GC shows promise as a method for rapid estimation of TPH in soil. A major hurdle in the evaluation of methodology for hydrocarbons residues in soil is the challenge posed by the weathering of such residues.     

Biotechnology and bioremediation: successes and limitations

With advances in biotechnology, bioremediation has become one of the most rapidly developing fields of environmental restoration, utilizing microorganisms to reduce the concentration and toxicity of various chemical pollutants, such as petroleum hydrocarbons, polycyclic aromatic hydrocarbons, polychlorinated biphenyls, phthalate esters, nitroaromatic compounds, industrial solvents, pesticides and metals. A number of bioremediation strategies have been developed to treat contaminated wastes and sites. Selecting the most appropriate strategy to treat a specific site can be guided by considering three basic principles: the amenability of the pollutant to biological transformation to less toxic products (biochemistry), the accessibility of the contaminant to microorganisms (bioavailability) and the opportunity for optimization of biological activity (bioactivity). Recent advances in the molecular genetics of biodegradation and studies on enzyme-tailoring and DNA-shuffling are discussed in this paper.     

Colorimetric method for a rapid detection of oxygenated aromatic biotransformation products

The quantitative production of the oxygenated products from the biotransformation of aromatic substrates can be detected using a very simple and rapid colorimetric test. The method is based on Gibbs' reagent (2,6-dichloroquinone-4-chloroimide) and has been developed for routine spectrophotometric or microtitre plate assay allowing the detection of products with a sensitivity as low as 5 mM.     

Metabolic fingerprinting of rat urine by LC/MS Part 1. Analysis by hydrophilic interaction liquid chromatography-electrospray ionization mass spectrometry

Complex biological samples, such as urine, contain a very large number of endogenous metabolites reflecting the metabolic state of an organism. Metabolite patterns can provide a comprehensive signature of the physiological state of an organism as well as insights into specific biochemical processes. Although the metabolites excreted in urine are commonly highly polar, the samples are generally analyzed using reversed-phase liquid chromatography mass spectrometry (RP-LC/MS). In Part 1 of this work, a method for detecting highly polar metabolites by hydrophilic interaction liquid chromatography-electrospray ionization mass spectrometry (HILIC/ESI-MS) is described as a complement to RP-LC/ESI-MS. In addition, in an accompanying paper (Part 2), different multivariate approaches to extracting information from the resulting complex data are described to enable metabolic fingerprints to be obtained. The coverage of the method for the screening of as many metabolites as possible is highly improved by analyzing the urine samples using both a C(18) column and a ZIC-HILIC column. The latter was found to be a good alternative when analyzing highly polar compounds, e.g., hydroxyproline and creatinine, to columns typically used for reversed-phase liquid chromatography.     

Bioremediation of oil contaminated beach sediments using indigenous microorganisms in singapore

The potential of using indigenous microorganisms in beach sediments to degrade petroleum hydrocarbons emanating from marine oil spillages in the Straits of Singapore was investigated. A field trial was conducted using oil contaminated beach sediments from Pulau Semakau – a small island 15 km south of Singapore. The results clearly show that the addition of inorganic nutrients to beach sediments significantly enhanced the activity of indigenous microorganisms (measured using the dehydrogenase enzyme assay and viable cell count techniques), as well as the removal of total recoverable petroleum hydrocarbons (TRPH) over a 50-day study period (with up to 44% in the case of nutrient addition). The potential of exploiting in-situ bioremediation techniques for oil spill clean-up operations in tropical marine environments is discussed.