Harold C. Thompson

Simultaneous determination of malachite green, gentian violet and their leuco metabolites in catfish or trout tissue by high-performance liquid chromatography with visible detection

A sensitive analytical procedure for the determination of residues of leucomalachite green (LMG)-malachite green (MG) and leucogentian violet (LGV)-gentian violet (GV) in catfish or trout tissue is presented. Frozen (-20 degrees C) fish fillets were cut into small pieces and blended in a Waring blender. A 20-g amount of homogenized fish tissue was extracted with acetonitrile-buffer, partitioned against methylene chloride, and cleaned up on tandem neutral alumina and propylsulfonic acid cation-exchange solid-phase extraction cartridges. Samples of 100 microliters (0.8 g equiv.) were chromatographed isocratically in 10 min using an acetonitrile-buffer mobile phase on a short-chain deactivated (SCD) reversed-phase column (250 x 4.6 mm I.D.) in-line with a post-column PbO2 oxidation reactor. The PbO2 post-column reactor efficiently oxidized LMG to the chromatic MG, and LGV to the chromatic GV permitting visible detection at 588 nm for all four compounds. Linearity was demonstrated with standards over the range of 0.5-50 ng per injection. Recoveries of LMG, MG, LGV and GV from catfish tissues fortified at 10 ng/g were 75.4 +/- 3.0, 61.3 +/- 4.1, 72.6 +/- 3.7 and 87.9 +/- 2.5, respectively, while trout tissues fortified at 10 ng/g yielded recoveries of 82.6 +/- 2.3, 48.6 +/- 1.8, 72.1 +/- 2.1 and 83.8 +/- 4.6 (mean +/- S.D., n = 4), respectively.

Persistence of gentian violet and leucogentian violet in channel catfish (ictalurus punctatus) muscle after water-borne exposure

Gentian violet is a triphenylmethane dye that is an antifungal/antiparastic agent. GV is similar to malachite green that has been used in the aquaculture industry for treatment or prevention of external fungal and parasitic infections in fish and fish eggs although it (MG) is not approved for this use. For these reasons, GVs potential for misuse by the aquaculture industry is high. The uptake and depletion of gentian violet (GV) were determined in channel catfish (Ictalurus punctatus) after water-borne exposure (100 ng ml(-1), 1 h) under simulated aquaculture farming conditions. Leucogentian violet (LGV) was rapidly formed, concentrated in the muscle tissue, and very slowly eliminated from muscle tissue. An isocratic (60% acetonitrile-40% water; 0.05 M ammonium acetate buffer, pH 4.5) HPLC system consisting of a 5 microm LC-CN 250x4.6 mm I.D. column, a 20x2.0 mm I.D. PbO2 oxidative post-column, and a UV-VIS detector set at 588 nm were used to determine uptake and depletion of tissue residues of GV and LGV with time. GV was rapidly depleted and converted to its major metabolite, LGV, which was detected out to 79 days. Therefore, LGV is the appropriate target analyte for monitoring exposure of channel catfish to GV.

SFE extraction of aflatoxins (B1, B2, G1, and G2) from corn and analysis by HPLC

A supercritical-fluid extraction (SFE) method has been developed that extracts aflatoxins (B1, B2, G1 and G2) from spiked corn using modified supercritical carbon dioxide. Methanol is added to the SFE extraction cell containing the corn layered between Hydromatrix (a dispersing material) which helps to prevent the clogging of the frits and reduces the effect of moisture on the extraction. The corn is held in static extraction at 65°C and 51.7 MPa for 15 min followed by a dynamic extraction with 20 mL of liquid carbon dioxide. The sample is depressurized and modifier re-added with a 10-min static extraction at 51.7 MPa followed by a dynamic extraction with 20 mL of liquid carbon dioxide. The SFE extract is collected in 10 mL of chloroform and further cleaned up with a Florisil Sep-Pak. The aflatoxins were analyzed by HPLC using fluorescence detection after post-column derivatization with iodine. Recoveries of the aflatoxins B1, B2, G1, and G2) over a range of 3 to 11 ng g-1 averaged 77.3, 82.9, 75.4, and 80.3%, respectively.

Determination of lincomycin residues in salmon tissues by gas chromatography with nitrogen-phosphorus detection

A sensitive method for the determination of lincomycin residues in fish tissues is described. Lincomycin was extracted from fish tissues with phosphate buffer (pH 4.5). The extract was concentrated with a C18 solid-phase extraction cartridge and further cleaned up by solvent extraction. Lincomycin was derivatized with N,O-bis(trimethylsilyl)trifluoroacetamide to form a trimethylsilyl derivative before being analyzed by gas chromatography with nitrogen-phosphorus detection. Coumaphos was used as the internal standard. Assays showed good linearity in the range 25-250 ppb (ng/g) (r = 0.9994). Recoveries of fortified lincomycin at 50, 100 and 200 ppb were > 80% with relative standard deviations < 6%. The limit of detection of the method was 1.7 ppb and the limit of quantitation was 3.8 ppb.

Determination of N-nitrosamines and N-nitrosamine precursors in rubber nipples from baby pacifiers by gas chromatography-thermal energy analysis

N-Nitrosamines and precursors are present in rubber products in which the accelerators and stabilizers used in the vulcanization process were derived from dialkylamines. Research was performed to develop data concerning the presence of N-nitrosamines and precursors so that the health significance of the exposure problem related to infant ingestion of these chemicals could be properly assessed. Volatile N-nitrosamines were determined in cut-up pacifier nipples by extraction with dichloromethane followed by concentration in a Kuderna-Danish evaporator, high-temperature mineral oil purge and trap, and analysis by gas chromatography--thermal energy analysis (GC-TEA). N-nitrosodibutylamine (NDBA) was the principal N-nitrosamine found, with concentrations up to 427 ppb. N-Nitrosamines and precursors in cut-up and intact nipples were determined by GC-TEA after a single extraction with artificial saliva. NDBA was the principal nitrosamine found, at levels up to 1040 ppb, while dibutylamine (DBA) was the principal precursor found, at levels up to 3890 ppb. The persistence of these compounds in intact nipples was determined by multiple artificial saliva extractions. Amounts of NDBA and DBA found after 15 artificial saliva extractions of intact pacifier nipples totalled 824 ppb and 15.6 ppm, respectively. N-Nitrosamine levels generally showed a gradual decrease in concentration with each extraction, whereas no consistent trend could be determined for concentrations of precursors.

Determination of leucogentian violet and gentian violet in catfish tissue by high-performance liquid chromatography with visible detection

A sensitive analytical procedure for the determination of residues of leucogentian violet (LGV) and gentian violet (GV) in catfish tissue is presented. Frozen (-20 degrees C) catfish fillets were cut into chunks and then blended in a Waring blender. A 10-g amount of catfish muscle tissue was homogenized and extracted with acetonitrile-buffer, partitioned against methylene chloride, and cleaned up on tandem neutral alumina and propylsulfonic acid cation-exchange solid-phase extraction cartridges. Samples of 100 microliters (0.5 g equiv.) were chromatographed isocratically in 15 min using an acetonitrile-buffer mobile phase on a cyano phase column in-line with a post-column PbO2 oxidation reactor. The PbO2 post-column reactor efficiently oxidized the LGV to the chromatic GV permitting visible detection at 588 nm for both LGV and GV. Linearity was demonstrated with standards over the range 0.5-50 ng per injection. Recoveries of LGV and GV from catfish tissues fortified at 20, 10, and 1 ng/g were 83.1 +/- 1.2, 78.4 +/- 4.0, 84 +/- 8 and 92.7 +/- 1.8, 95.0 +/- 2.2, 93 +/- 2 (mean +/- S.D., n = 4), respectively.

Trace analysis of the antihistamines methapyrilene hydrochloride, pyrilamine maleate and triprolidine hydrochloride monohydrate in animal feed, human urine and wastewater by high-performance liquid chromatography and gas chromatography with nitrogen-phosphorus detection.

Toxicological evaluation of the antihistamines methapyrilene hydrochloride, pyrilamine maleate, and triprolidine hydrochloride monohydrate using methapyrilene hydrochloride as the positive indicator was investigated as part of a structure-activity relationship study in rats and mice. Prerequisites for the toxicological tests were the development of analytical procedures to certify the dose, homogeneity and stability of the drugs in animal feed and to monitor human urine for possible exposure and to ensure removal of the test agents from wastewater prior to its discharge into the environment. A high-performance liquid chromatographic (HPLC) system was developed using a fluorescence detector for the determination of methapyrilene hydrochloride and pyrilamine maleate in feed at levels as low as 100 ng/g and in human urine as low as 1 ng/g. An HPLC-UV procedure was developed for the determination of triprolidine hydrochloride monohydrate in feed at levels as low as 10 micrograms/g. Data concerning p-values, extraction efficiencies from feed and stability experiments in feed are presented for these antihistamines. A gas chromatographic procedure using a nitrogen-phosphorus detector was also developed for determining the three antihistamines in admixture in wastewater at levels as low as 10 ng/g.

High-performance liquid chromatographic analysis of the antituberculosis drugs aconiazide and isoniazid

Reversed-phase HPLC methods are described for determining the stability and concentration certification of the antituberculosis prodrug aconiazide (ACON) in aqueous dosing solution and for assessing the concentrations of ACON and isoniazid (INH) in plasma from ACON-treated male and female Fischer-344 rats. ACON was analyzed in plasma by direct injection; it was separated on a 250 x 4.6 mm I.D. 5 microns C18 column using a 40% aqueous methanol mobile phase containing 5 g/l ammonium formate, and detected at 313 nm. INH was determined in the plasma of treated rats after a two-step precipitation of plasma proteins; it was separated on a 250 mm x 4.6 mm I.D. 5 microns CN column, eluted with 5% aqueous isopropanol containing 5 g/l ammonium formate, and detected with an electrochemical detector at +0.8 V. These methods allow a simple, rapid, and reliable determination of ACON and INH in plasma down to 0.1 micrograms/ml.

Application of the particle beam interface to high-performance liquid chromatography-thermal energy analysis and electron impact mass spectrometry for detection of non-volatile N-nitrosamines

Abstract Interest in the analysis of non-volatile N-nitrosamines has recently been renewed due to the development of several new reversed-phase HPLC interfaces to thermal energy analysis (TEA) or chemiluminescence detection. A new application of a counter flow gas diffusion cell (CFGDC)-based particle beam LC interface (Universal Interface, Vestec) is described for the HPLC-TEA analysis of the non-volatile N-nitrosamines, N-Nitrosodiethanolamine (NDELA) and N-nitrosomethyl-p-amino-2-ethylhexylbenzoate (NMPABAO). The interface incorporates a thermospray vaporizer, desolvation chamber, and CFGDC to reduce the LC effluent to a dry aerosol and a single-stage momentum separator to form a particle beam of the non-volatile analyte. Using this system, the LC-TEA response to NDELA was linear in the range 6–200 ng total amount injected. Several experiments are reported indicating the effect of thermospray tip temperature, He carrier flow-rate, and mobile phase composition on TEA response. Minimum detection limits (5 ng NDELA injected on column) are comparable to other LC-TEA interfacing methods. Several advantages over existing methodology which include ease of use, ruggedness and MS compatibility are discussed. Additional LC-particle beam MS data are reported indicating that full scan electron impact MS identification of the N-nitrosamine contaminants in cosmetics is possible for confirming TEA detection data.

Impact of membrane desolvation on the effects of sodium on response in inductively coupled plasma atomic emission spectrometry with ultrasonic sample introduction

This report compares the influence of sodium on response with ICP-AES for copper, cadmium, lead and cobalt with ultrasonic sample introduction in the presence and absence of a microporous membrane desolvation (MMD). Comparisons are reported at viewing positions of 8, 14, and 20 mm above the load coil, for injector gas flow rates of 0.70, 0.85 and 1.00 l min–1, and at sodium concentrations ranging from 0.00 to 0.08 M Na. Blank emission intensities were depressed in the presence of sodium regardless of the operating condition(s). Depressions in the blank emission intensities were greater without the MMD, but were less than 20%. The magnitude of depression in the blank emission intensities did not vary significantly among the analyte lines or with the viewing position. The impact of sodium on signal intensities with and without the MMD was greater than the impact on the blank emission intensities, with changes approaching 150%. At the lower viewing positions, signal enhancement in the presence of sodium was common and the degree of enhancement was greater with the MMD. Response changes with respect to sodium concentration were also different with and without the MMD low in the plasma. Higher in the plasma, element-to-element variation occurred in the nature of the impact of sodium. For signals that were enhanced, the degree of enhancement was generally greater with the MMD, but when signals were depressed, there was no correlation to the presence or absence of the MMD. In contrast to the lower viewing position, higher in the plasma response changes with respect to the sodium concentration were similar. These results suggest that although the plasma is physically different with and without the MMD, the MMD does not alter the mechanism(s) by which sodium impacts response.