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Nitisinone (2-(2-nitro-4-trifluoromethylbenzoyl)-1,3-cyclohexanedione, NTBC) was the first synthetically produced triketone herbicide. However, its unsatisfactory herbicidal properties, negative impact on the natural environment and the high cost of synthesis have hindered its commercialization as a plant protection agent. Nevertheless, NTBC has become the medical treatment of choice for a rare hereditary metabolic disease -hepatorenal tyrosinemia. Literature review shows that most research on nitisinone focuses on its medical applications, while there are neither in-depth studies of its stability nor its degradation pathways. Therefore, the aim of our study was to employ liquid chromatography coupled with mass spectrometry (LC-MS/MS) to determine the stability of NTBC in different experimental conditions (pH of solution, temperature, time of incubation, ultraviolet radiation), identify its degradation products and determine the stability of the latter. Electrospray ionization (ESI) in the negative ion mode was used as an ionization method and the analytes were detected by multiple reaction monitoring. We show that nitisinone stability increases with increasing pH of the solution. At pH similar to that of gastric juice in the human stomach, two major products of NTBC degradation are formed: 2-amino-4-(trifluoromethyl)benzoic acid (ATFA) and 2-nitro-4-(trifluoromethyl)benzoic acid (NTFA), which show considerable stability under studied conditions. The results of these studies shed new light on the properties of NTBC, therefore contributing to better understanding of possible risks and benefits of its medical application., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Most solid tumors contain cancer-associated fibroblasts (CAFs) that support tumorigenesis and malignant progression. However, the cellular origins of CAFs in epithelial ovarian cancers (EOCs) remain poorly understood, and their utility as a source of clinical biomarkers for cancer diagnosis has not been explored in great depth. Here, we report establishing in vitro and in vivo models of CAFs in ovarian cancer development. Normal ovarian fibroblasts and mesenchymal stem cells cultured in the presence of EOC cells acquired a CAF-like phenotype, and promoted EOC cell migration in vitro. CAFs also promoted ovarian cancer growth in vivo in both subcutaneous and intraperitoneal murine xenograft assays. Molecular profiling of CAFs identified gene expression signatures that were highly enriched for extracellular and secreted proteins. We identified novel candidate CAF-specific biomarkers for ovarian cancer including NPPB, which was expressed in the stroma of 60% primary ovarian cancer tissues (n = 145) but not in the stroma of normal ovaries (n = 4). NPPB is a secreted protein that was also elevated in the blood of 50% of women with ovarian cancer (n = 8). Taken together, these data suggest that the tumor stroma is a novel source of biomarkers, including NPPB, that may be of clinical utility for detection of EOC., (© 2014 UICC.)

The immobilization of microorganisms is essential for efficient bioaugmentation systems. The performance of Bacillus cereus G5 as biofilm-forming bacteria and Comamonas testosteroni A3 a 3,5 dinitrobenzoic acid (DNB)-degrading strain] in laboratory-scale sequencing batch biofilm reactors (SBBRs) treating DNB synthetic wastewater has been examined. The microbial diversity in the reactors was also explored. The reactor R3 inoculated with B. cereus G5 and C. testosteroni A3 together not only improved the removal of contaminants, but also exhibited obvious resistance to shock loading with DNB during later operations. Pyrosequencing was used to evaluate bacterial communities in three reactors. Comamonas was predominant in the reactor R3, indicating the effect of G5 in promoting immobilization of A3 cells in biofilms. Those microbial resources, e.g.G5, which can stimulate the self-immobilization of the degrading bacteria offer a novel strategy for immobilization of degraders in bioaugmentation systems and show broader application prospects.

The nitroaromatic compound 4-bromomethyl-3-nitrobenzoic acid (ANB) is a promising antitumoral agent whose activity has recently been investigated. Forced degradation studies were conducted on ANB with a high-performance liquid chromatography-ultraviolet assay to establish its stability and selectivity. ANB was subjected to degradation studies under hydrolytic (acid and alkaline), oxidative and light exposition conditions. The compound showed greater lability only in acid and alkaline conditions by forming a major degradation product. The chromatographic separation of ANB and its degradation product was achieved on an octadecylsilane column using a mobile phase of methanol-water (80:20, v/v), pH 4.0 adjusted with formic acid, flow rate of 0.7 mL/min, ultraviolet diode array detection at λ 271 nm, injection volume of 20 µL and temperature of 30°C. The method was validated according to International Conference on Harmonization guidelines with respect to precision (average relative standard deviation 0.67%), accuracy (average 99.97%), linearity (y = 118730x + 12912; coefficient of determination > 0.999), specificity and robustness (p > 0.05). The degradation product formed as a result from the hydrolysis of nitrobenzyl bromide, corresponded to its benzyl alcohol, 4-hydroxymethyl-3-nitrobenzoic acid (ANOH) and was characterized by co-elution with a synthetic ANOH working standard., (© The Author [2013]. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Three legal highs; nitracaine (3-(diethylamino)-2,2-dimethylpropyl 4-nitrobenzoate), methoxypiperamide (MEOP, (4-methoxyphenyl)(4-methylpiperazin-1-yl)methanone) and mephtetramine (MTTA, 2-((methylamino)methyl)-3,4-dihydronaphthalen-1(2H)-one) appeared in 2013 as new psychoactive substances (NPS) on Internet websites selling 'research chemicals'. These compounds were synthesized and analyzed via our synthesize, analyze, and metabolize (SAM) protocol. Nitracaine was synthesized by the transesterification of methyl 4-nitrobenzoate with 3-(diethylamino)-2,2-dimethylpropan-1-ol. Methoxypiperamide was synthesized by the reaction of 4-methoxybenzoyl chloride with 1-methylpiperazine, and mephtetramine through the Mannich reaction of 1-tetralone with paraformaldehyde and methylamine hydrochloride. Each compound was characterized by nuclear magnetic resonance (NMR), gas chromatography with electron impact mass spectrometry (GC-EIMS), liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS), and high resolution electrospray ionization mass spectrometry (HR-ESI-MS). A sample of nitracaine was also test-purchased from an Internet vendor and its structure confirmed by GC-EIMS and LC-ESI-MS. Finally, the in vitro metabolism of the nitracaine, mephtetramine, and methoxypiperamide was investigated, using a human microsomal liver extract, in order to tentatively identify potential metabolites that may be encountered in the analysis of biological samples in clinical or toxicology labs. The use of our SAM protocol highlights the ability of academic research labs to quickly respond to and disseminate information about emerging NPS., (Copyright © 2014 John Wiley & Sons, Ltd.)

A rapid, highly selective, and sensitive method was developed for detecting fluoroglycofen-ethyl in soybean seed, plant, and soil using UPLC-MS-MS. The detection limits of fluoroglycofen-ethyl in soybean seed, plant, and soil were 0.5, 1, and 1 μg kg(-1), respectively. Recoveries ranged from 83.4% to 99.2%, in which intra-day RSDs were from 1.3% to 6.7% and inter-day RSDs were from 1.9% to 7.0%. In the dissipation study, the half-lives of fluoroglycofen-ethyl were 34.8 (Shanxi) and 48.5 h (Heilongjiang) in soil and 43.3 h in soybean plant in both locations. The residues of fluoroglycofen-ethyl in all samples were below LODs 30 days before and during harvest.

The detection of nitroaromatic compounds, best known as raw materials in explosives preparations, is important in many fields including environmental science, public security and forensics. CdSe quantum dots capped with PAMAM-G(4) dendrimer were synthetized in water and used for the detection of trace amounts of three nitroaromatic compounds: 4-methoxy-2-nitrophenol (MNP), 2-amine-5-chloro-1,3-dinitrobenzene (ACNB) and 3-methoxy-4-nitrobenzoic acid (MNB). To increase the apparent water solubility of these compounds α-cyclodextrin (α-CD) was used to promote the formation of inclusion complexes. The studied nitroaromatic compounds (plus α-CD) significantly quenched the fluorescence intensity of the nanocomposite with linear Stern-Volmer plots. The Stern-Volmer constants (standard deviation in parenthesis) were: MNB, K(SV)=65(5)×10(4) M(-1); ACNB, K(SV)=19(2)×10(4) M(-1); and, MNP, K(SV)=33(1)×10(2) M(-1). These constants suggest the formation of a ground state complex between the nitroaromatric compounds and the sensor which confers a relatively high analytical sensitivity. The detection sensibilities are about 0.01 mg L(-1) for MNB and ACNB and about 0.1 mg L(-1) for MNP. No interferences or small interferences are observed for trinitrotoluene [K(SV)=10(2)×10(2)×M(-1)], 2,4-dinitrotoluene [K(SV)=20(3)×10 M(-1)], 2,6-dinitrotoluene [K(SV)=11(4)×10 M(-1)] and nitrobenzene [K(SV)=2(1)×10(3)×M(-1)]., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Many pesticides in use are chiral compounds containing stereoisomers. However, the environmental behavior and fate of such compounds with respect to enantioselectivity so far has received little attention. In this study, the degradation of lactofen and its main metabolites (acifluorfen, an achiral compound; desethyl lactofen, a chiral compound) in sediment were investigated under laboratory conditions using enantioselective HPLC, and the enantioselectivities of individual enantiomers of lactofen and desethyl lactofen in acute toxicity to Daphnia magna were studied. The calculated LC(50) values of S-(+)-, rac-, and R-(-)-lactofen were 17.689, 4.308, and 0.378 microg/mL, respectively, and the calculated LC(50) values of S-(+)-, rac-, and R-(-)-desethyl lactofen were 21.327, 13.684, and 2.568 mug/mL, respectively. Therefore, the acute toxicities of lactofen and desethyl lactofen enantiomers were enantioselective. In sediments, S-(+)-lactofen or S-(+)-desethyl lactofen was preferentially degraded, resulting in relative enrichment of the R-(-)-form. Lactofen and desethyl lactofen were both configurationally stable in sediment, showing no interconversion of S- to R-enantiomers or vice versa. Furthermore, the conversion of lactofen to desethyl lactofen proceeded with retention of configuration. These results for major differences in acute toxicity and degradation of the enantiomers may have some implications for better environmental and ecological risk assessment for chiral pesticides.

Objective: To determine residues of multiple biphenyl ether herbicides simultaneously in water using high performance liquid chromatography (HPLC) with cloud-point extraction., Methods: The residues of eight biphenyl ether herbicides (including bentazone, fomesafen, acifluorfen, aclonifen, bifenox, fluoroglycofenethy, nitrofen, oxyfluorfen) in water samples were extracted with cloud-point extraction of Triton X-114. The analytes were separated and determined using reverse phase HPLC with ultraviolet detector at 300 nm. Optimized conditions for the pretreatment of water samples and the parameters of chromatographic separation applied., Results: There was a good linear correlation between the concentration and the peak area of the analytes in the range of 0.05-2.00 mg/L (r = 0.9991-0.9998). Except bentazone, the spiked recoveries of the biphenyl ether herbicides in the water samples ranged from 80.1% to 100.9%, with relative standard deviations ranging from 2.70% to 6.40%. The detection limit of the method ranged from 0.10 microg/L to 0.50 microg/L., Conclusion: The proposed method is simple, rapid and sensitive, and can meet the requirements of determination of multiple biphenyl ether herbicides simultaneously in natural waters.

The FTIR and FT-Raman spectra of 5-amino-2-nitrobenzoic acid (ANB) have been recorded in the region 400-4000 cm(-1). The observed frequencies were assigned to different modes of vibrations on the basis of fundamental, combination and overtones. The geometry has been optimized with complete relaxation on the potential energy surface at HF, MP2 and B3LYP level of theories using 6-311++G(d,p) basis set and compared with the crystal data. The possible hydrogen bond interaction has been estimated taking a model compound. Further harmonic vibrational frequency calculations have been carried out at HF and B3LYP levels and the scaled values were in good agreement with majority of the experimental observations. The theoretically constructed spectra coincide satisfactorily with those of experimental spectra.

The sensitivity of the Slusher and Hahn's nuclear quadrupole double resonance technique is calculated in general for an arbitrary nuclear spin S of the quadrupole nuclei and for an arbitrary asymmetry parameter eta of the electric field gradient tensor. The nuclear spin S=5/2 ((17)O, (25)Mg, ...) is treated in details. The influence of the cross-relaxation rate between the quadrupole nuclei and the abundant spin system on the sensitivity of double resonance is discussed. The results of the theoretical analysis are applied in the analysis of the (1)H-(17)O nuclear quadrupole double resonance spectra in p-toluenesulfonamide and 2-nitrobenzoic acid. The 17O nuclear quadrupole resonance frequencies from a sulfonamide group are determined for the first time. The proton-oxygen cross-relaxation rates and the proton local frequency in zero external magnetic field are experimentally determined from the nuclear quadrupole double resonance spectra.

Five groundwater samples from the former ammunition production site at Elsnig, Germany, were analyzed for highly polar components by LC-NMR and LC-MS. A variety of unknown pollutants could be identified. Possibilities and limitations of the combined use of LC-NMR and LC-MS techniques for on-line identification are discussed. Further unknown components were identified through isolation by HPLC cuts and off-line NMR and MS investigations. Most of the polar compounds in the investigated samples could also be quantified., (Copyright 2005 John Wiley & Sons, Ltd.)

A method is described for determination of the steroidal saponin, eruboside B, originating in garlic and garlic products as the p-nitrobenzoyl chloride (PNBC) derivative by reversed-phase liquid chromatography (with ultraviolet detection at 260 nm. Proto-eruboside B was extracted from garlic (Allium sativum L.); subjected to solid-phase extraction (SPE) with a C18 cartridge, Florisil column chromatography, and silica gel column chromatography; and then enzymatically converted to eruboside B, which was applied as an external standard. Steroidal saponins in garlic and commercial garlic products were extracted with methanol and purified by SPE cartridges, followed by enzymatic treatment. A frostanol saponin such as proto-eruboside B is enzymatically transformed to a spirostanol saponin, eruboside B. After the derivatization with PNBC, the saponin derivative was chromatographed on a C8 column with a gradient elution of (A) 80% aqueous acetonitrile and (B) 100% acetonitrile. The detection limit of the developed method was 1 microg/g for the samples. The method was applied to the analysis of garlic and garlic health food products available in Japan.

The problem of maintaining the moisture content of samples throughout the course of a soil photolysis study is addressed. The photolytic degradations of asulam, triclopyr, acifluorfen, and atrazine were independently compared in air-dried soils and in moist (75% field moisture capacity at 0.33 bar) soils maintained at initial conditions through the use of a specially designed soil photolysis apparatus. Each pesticide was applied at 5 microg/g. The exposure phase extended from 144 to 360 h, depending on the half-life of the compound. A dark control study, also using moist and air-dried soils, was performed concurrently at 25 degrees C. The results showed significant differences in half-life. The dissipations generally demonstrated a strong dependence on moisture. In most cases, photolytic degradation on air-dried soil was longer than in the moist dark control soils. Half-lives in dry soil were 2-7 times longer, and in the case of atrazine, the absence of moisture precluded significant degradation. Moist soil experiments also tended to correlate more strongly with linear first-order degradations. The dark control experiments also demonstrated shorter half-lives in moist soil. Moisture was also observed to affect the amount of degradate formed in the soils.

Direct liquid chromatographic separations of the enantiomers of metoprolol and bisoprolol have been developed, using (R)-1-naphthylglycine and 3,5-dinitrobenzoic acid as chiral stationary phase (CSP). The separations were achieved in a normal phase system employing a mobile phase containing n-hexane, 1,2-dichloroethane and methanol. Column efficiency was strongly dependent on the composition of the mobile phase. The eluent contents of methanol and of 1,2-dichloroethane were optimized, and so was flow-rate and column temperature. Under the optimal conditions, linear responses for (R)-metoprolol and (S)-metoprolol are obtained in the range of 0.079-1.38 and 0.015-5.80 mg/ml, with detection limits of 0.008 and 0.002 mg/ml, respectively. As for bisoprolol, the linear ranges of (R)-isomer and (S)-isomer are 0.05-1.31 and 0.02-1.00 mg/ml with detection limits of 0.001 and 0.008 mg/ml, respectively. The relative standard deviation (R.S.D.) of each enantiomer did not exceed 0.90%. The method has been successfully applied to the determination of enantiomers in pharmaceuticals.

We show that anthocyanins can function as potent inhibitors of the formation of nitrated tyrosine in vitro, and clarified how pelargonidin (Pel), which has a mono-hydroxyl group on the B-ring, can scavenge peroxynitrite (ONOO(-)) by detection of the reaction products. Pel was reacted with ONOO(-), then the reaction mixture was analyzed using high-performance liquid chromatography (HPLC). The HPLC analyses showed two novel peaks assumed to be the reaction products. Based on the instrumental analyses, the reaction products were identified as p-hydroxybenzoic acid and 4-hydroxy-3-nitrobenzoic acid. Pel can protect tyrosine from undergoing nitration through the formation of p-hydroxybenzoic acid and 4-hydroxy-3-nitrobenzoic acid.

Nitrotoluene and nitrobenzoic acid isomers were separated completely in less than 10 minutes by Kromasil C18 (200 mm x 4.6 mm i.d.), when V(CH3OH):V(H2O):V(THF) = 55:44:1 with beta-CD of 0.02 mol/L at flow rate of 2.0 mL/min (former 4 min) and 2.6 mL/min (later 6 min). Detector of UV-254 at 0.04 AUFS was used.

An analytical method based on the use of a liquid chromatograph equipped with a UV detector was developed for the determination of acifluorfen in soil and water. Acifluorfen was extracted from soil in methanol-0.10N NaOH (80 + 20 v/v) and from water by partition with dichloromethane. Solvent partitioning and solid-phase extraction were used to separate acifluorfen from major interfering sample components. Average recoveries from soil at 1, 0.1, and 0.01 ppm fortification levels were 95.1 +/- 3.4, 92.6 +/- 2.9, and 73.9 +/- 3.0%, respectively. Recoveries from water spiked at levels from 0.01 to 1 ppm averaged 96.5 +/- 5.4%. Method limits of detection were 0.006 ppm in soil and 0.003 ppm in water.

The direct separation of 26 bay region and non-bay region mono-ol and diol enantiomers of phenanthrene, benz[a]anthracene, and chrysene was compared by high-performance liquid chromatography on commercially available columns, packed with gamma-aminopropylsilanized silica to which either (R)-N-(3,5-dinitrobenzoyl)phenylglycine(R-DNBPG) or (S)-N-(3,5-dinitrobenzoyl)leucine(S-DNBL) was either ionically or covalently bonded. In general, enantiomers of bay region mono-ols and diols are more efficiently resolved than those of non-bay region derivatives. Elution orders of enantiomers on either chiral stationary phase are the same, regardless of whether the chiral stationary phase is ionically or covalently bonded. Except for the enantiomers of 4-hydroxy-4-methyl-1,2,3,4-tetrahydrobenz[a]anthracene, 1,2,3,4-tetrahydrobenz[a]anthracene trans-1,2-diol, and benz[a]anthracene trans-1,2-dihydrodiol, elution orders of resolved enantiomers on R-DNBPG are reversed on S-DNBL. The enantiomers are generally more efficiently resolved on R-DNBPG than on S-DNBL. With the exception of the elution order of the enantiomeric 4-hydroxy-1,2,3,4-tetrahydrochrysene, the results of this study are consistent with the chiral recognition mechanisms proposed by Pirkle and co-workers, who developed the chiral stationary phases used in this study.

The rapid resolution of racemic N-4-nitrobenzoylamino acid isopropyl esters was accomplished without the loss of enantioselectivity by supercritical fluid chromatography (SFC) on novel chiral valine-diamide phases with carbon dioxide and a polar methanol modifier. In each stationary phase, a chiral moiety was anchored to the silica gel surface by a long decamethylene spacer. The enantioselectivity in SFC was comparable to that in liquid chromatography using 2-propanol-n-hexane. The time required for analysis was less than 5 min, and the range of enantiomer resolution (Rs) was 10.8-1.25. On using 2-propanol in place of methanol the separation was improved, but was accompanied by a decrease in column efficiency. The end-capping effect of the remaining surface silanols on enantiomer resolution is discussed.

Sigma S comprises both free thiol groups and labile S-S-bonds of the total IgG that react with dithionitrobenzoate. Previous investigations have shown that this parameter was significantly decreased in malignant diseases of various organ systems as compared with benign disorders of the respective organs. The present study on patients with ovarian cancer investigated whether a significant correlation exists between the post-operative course of sigma S and the clinical evidence. For this purpose, on the basis of former results, a value of sigma S of 1.25 was determined as a criterion between malignant and benign ovarian diseases. For each investigated case, the post-operative temporal average of sigma S was calculated (sigma S) and a value of 1.25 was used as discriminating threshold. A four-fold table was set up containing the frequencies of tumor recurrences and tumor free cases with sigma S less or greater than 1.25. In a total of 40 patients of ovarian cancer an association between sigma S less than 1.25 and increased frequency of recurrences was confirmed, with a probability of error of 1%. The total predictive value of the sigma S level was about 70% and is herewith comparable to the leading tumor marker CA 125 in ovarian cancer.

Recently an elegant method for the quantification of the number of disulfide bonds in proteins and peptides has been reported [T.W. Thannhauser, Y. Konishi, and H.A. Scheraga (1984) Anal. Biochem. 138, 181-188]. The method is based on the quantification of 2-nitro-5-thiobenzoate (NTB) formed from the reaction of 2-nitro-5-thiosulfobenzoate with disulfides in the presence of excess sodium sulfite. Here it is reported that the NTB anion undergoes photochemical reaction with excess sulfite in the system, which results in the rapid disappearance of absorbance at 412 nm in the presence of light. The nonchromophoric derivative of this photochemical reaction is tentatively identified as a sulfo derivative of NTB. Based on these observations it is suggested that, for the quantification of disulfide bonds using the NTSB method, the assay should be carried out in the dark.

A method for the determination of cyanate concentration in blood plasma over the range 1 to 1000 microM is presented. Cyanate present in the dried residue of acetone-deproteinized plasma is converted to a chromophoric thiocarbamyl derivative by addition of pH 3.0-buffered thionitrobenzoic acid. The derivative is then analyzed by reversed-phase high-performance liquid chromatography with detection at 313 nm, near the absorption maximum. Carbamyl thionitrobenzoic acid peak height is quantified by comparison to a standard curve made by analysis of plasma samples to which known quantities of cyanate have been added. This technique is sensitive and linear with respect to cyanate concentration, and is faster than other reported methods; sample analysis and column regeneration are accomplished within 20 min.

The separation and quantitative determination of digitalis glycosides by high performance liquid chromatography following derivatization with 4-nitrobenzoylchloride (4-NBC1) is described. The compounds of primary interest were the digitalis glycosides and aglycones of the pharmaceutically important A, B and C series, The derivatization step results in higher extinction values at a more favourable wavelength (260 nm), which permits the use of low-cost ultraviolet detectors. Detection limits are below 20 ng/ml for all of the glycosides tested. The chromatographic properties are also improved by reducing the polarity without a decrease in selectivity. The use of low-polarity and low-viscosity solvent systems on silica gel adsorbents permits rapid isocratic separations of complex mixtures as they usually occur in pharmaceutical products and extracts. The quantitative potential of this method was demonstrated by analyzing ampoule solutions containing desacetyl lanatoside C as the active compound. The active substance, by-products and degradation products were determined down to 0.1% of the total glycoside concentration in one ampoule.

The direct resolution of K-region cis- and trans-dihydrodiol enantiomers of 14 unsubstituted and methyl- and bromo-substituted benz[a]anthracene (BA) derivatives was investigated by high-performance liquid chromatography with commercially available columns, packed with gamma-aminopropylsilanized silica to which either (R)-N-(3,5-dinitrobenzoyl)phenylglycine (R-DNBPG) or (S)-N-(3,5-dinitrobenzoyl)leucine (S-DNBL) is either ionically or covalently bonded. BA derivatives used in this study include: BA, 1-methyl-BA, 4-methyl-BA, 7-methyl-BA, 8-methyl-BA, 10-methyl-BA, 11-methyl-BA, 12-methyl-BA, 7,12-dimethyl-BA, 7-bromo-BA, 7-bromo-1-methyl-BA, 7-bromo-11-methyl-BA, 7-bromo-12-methyl-BA, and 3-methylcholanthrene. The enantiomers of BA trans-5,6-dihydrodiol were the only compounds not resolved by any of the four chiral stationary phases (CSPs) tested. The results indicate that conformational preference of the hydroxyl group is one of the most important factor in determining the elution order of dihydrodiol enantiomers. The presence and the location of a substituent and the molecular size and shape of the dihydrodiols can significantly affect the efficiency of enantiomeric resolution. In general, the ionically bonded R-DNBPG provides the best resolution of enantiomeric quasidiequatorial trans-dihydrodiols and the R,R enantiomers are consistently more strongly retained. In contrast, the enantiomeric pairs of quasidiaxial trans-dihydrodiols are generally better resolved by the covalently bonded R-DNBPG, and the S,S enantiomers are more strongly retained. The enantiomers of cis-dihydrodiols having hydroxyl groups that adopt quasiequatorial-quasiaxial and/or quasiaxial-quasiequatorial conformations are more consistently resolved by the ionically bonded S-DNBL and in all cases the S,R enantiomers are more strongly retained. Thus, it is possible to choose a CSP which resolves the K-region dihydrodiol enantiomers with a predictable elution order.

The use of electron capture negative ion desorption chemical ionization mass spectrometry was demonstrated in the analysis of phospholipid molecular species at the 1,3-dinitrobenzoate (DNB) diglyceride derivative. Modification of phosphatidylcholine (PC) or phosphatidylethanolamine (PE) by phospholipase C treatment and acylation of the resultant diglyceride with 3,5-dinitrobenzoylchloride afforded separation of the alkylacyl, alkenylacyl, and diacyl dinitrobenzoate subclasses by thin-layer chromatography (TLC). Separation of alkylacyl DNB into individual molecular species by reverse-phase high-performance liquid chromatography (RPHPLC) was demonstrated. Electron capture desorption chemical ionization of individual molecular species (10-25 ng) from a direct probe yielded a mass spectrum characterized by an intense molecular anion. This molecular anion was the base peak of the spectrum accounting for greater than 80% of the total ionization. From this molecular anion the total carbon number and degree of unsaturation of the fatty chains could be determined. Analysis of fatty acid content of the molecular species allowed unequivocal assignment of structure for the alkyl ether phospholipids. Using selected ion monitoring as little as 0.5 pmol of these species could be detected with a signal-to-noise ratio greater than or equal to 3. This technique was useful in the analysis of low picomolar amounts of molecular species of ether phospholipids in the rat lung. Given an appropriate internal standard, analysis of dynamic changes in turnover, metabolism and precursor product relationships could be undertaken.

Ion-pair reverse phase liquid chromatography (LC) and UV detection at 280 nm have been used to determine sodium acifluorfen (sodium-5-[2-chloro-4-(trifluoromethyl)-phenoxy]-2-nitrobenzoate), an experimental diphenyl ether herbicide, in dog feed. Sodium-5-(2,4-dichlorophenoxy)-2-nitrobenzoate is used as the internal standard. The feed is homogenized in 0.01N HCl, followed by ethyl acetate extraction, and centrifugation. The organic layer is removed and evaporated and the residue is reconstituted in methanol and filtered before LC analysis (mobile phase methanol-water (58 + 42), 0.005M in tetrabutylammonium phosphate and 0.045M in (NH4)2HPO4, at pH 7.4). The ion-pair technique offers a high degree of control over the retention characteristics of the herbicide and internal standard. The use of the internal standard permits precise and accurate quantitation and substantially reduces analysis time compared with the external standard method.

Residues of sodium 5-[2-chloro-4-(trifluoromethyl)phenoxy]-2-nitrobenzoate and its free acid are determined by treating a sample extract with diazomethane to convert the residues to methyl 5-[2-chloro-4-(trifluoromethyl)phenoxy]-2-nitrobenzoate. This compound is purified by chromatography on Florisil and measured by electron capture gas-liquid chromatography. The method has been used on soybeans, soybean foliage, soil, milk, and liver. It can detect 0.01 ppm with recoveries of 70--75%.