High-performance liquid chromatography of gentian violet, its demethylated metabolites, leucogentian violet and methylene blue with electrochemical detection

High-performance liquid chromatographic conditions are reported for the electrochemical detection (ED) of Gentian Violet, its demethylated metabolites, Leucogentian Violet and Methylene Blue. Gentian Violet, its demethylated metabolites and Leucogentian Violet were separated within 14 min on a cyano column eluted isocratically with methanol-buffer (60:40) as the mobile phase. ED responses for Gentian Violet, Leucogentian Violet and Methylene Blue were linear over the ranges 0.54-6.75, 0.50-25.2, and 5.7-285 ng, respectively. Under these conditions, the compounds were eluted in the following order: Leucogentian Violet, N"-2-tetra-methylparaosaniline chloride, N'-1-tetramethylpararosaniline chloride, pentamethylpararosaniline chloride and Gentian Violet. Methylene Blue and Gentian Violet had essentially the same retention time under these parameters. The detection limit for Gentian Violet, its demethylated metabolites and Leucogentian Violet was determined to be 0.1 pmol. A detection limit of 3 pmol was established for Methylene Blue. Detector response, elution, separation, linearity and sensitivity of detection are discussed.     

Determination and confirmation of malachite green and leucomalachite green residues in salmon using liquid chromatography/mass spectrometry with no-discharge atmospheric pressure chemical ionization

A liquid chromatography/mass spectrometry (LC/MS) method was developed to quantitate and confirm residues of leucomalachite green (LMG) in salmon tissue after their conversion to chromic malachite green (MG) in the extraction process. The method uses no-discharge atmospheric pressure chemical ionization (APCI) in conjunction with an ion-trap instrument to generate product-ion spectra. In the sample preparation procedure, salmon tissue is extracted with acetonitrile/buffer, the LMG residue is partitioned into methylene chloride, the LMG is converted to MG using an organic oxidizing agent, and the MG is isolated on alumina/propylsulfonic acid solid-phase extraction cartridges. The method was validated by fortifying salmon with different levels of LMG, and then detecting the residue as MG The LC/MS conditions, including a comparison of electrospray and no-discharge APCI, were evaluated and optimized. MG was not confirmed in any of the control tissue extracts, and all fortified samples analyzed during validation met the confirmation criteria as described. In addition to providing confirmatory data, this method can provide an alternative method for quantitation of MG in salmon. The recoveries of LMG measured as MG by this LC/MS method, at fortification levels of 1-10 ng/g were very high (86-109%), with low relative standard deviation(RSD) values (6.4-13%). The results agreed very closely with those obtained for the same extracts using an LCNIS procedure, indicating that matrix suppression was not an issue. The presence of LMG in salmon tissue samples fortified at 0.25 ng/g was confirmed by this method, with an average recovery of 70.1% and an RSD of 12.0%. Sample extracts from fish exposed to MG were also analyzed.     

Liquid chromatographic determination of diminazene diaceturate (Berenil) in raw bovine milk

A simple liquid chromatographic (LC) method is presented for the determination of diminazene (DZ) in raw bovine milk. DZ is extracted from raw milk by chilled aqueous centrifugation and is isolated from milk components on a cyano solid-phase extraction column. DZ is eluted by using a methanol-ion pairing reagent. A Phenomenex LUNA CN column and an acetonitrile-buffered mobile phase with a counter ion are used for gradient LC. The LC effluent is monitored at a detection wavelength of 372 nm by using a deuterium lamp. Under the parameters described, the retention time of DZ is 8-10 min with a peak area response of 6.5 mAU/ng. The method demonstrated excellent precision over all levels tested (25-400 ppb) with an overall average recovery of 90.4 +/- 14.5%. The method is applicable to the monitoring of milk for DZ residues at the 25 ppb level with a limit of quantitation of 10 ppb.     

No-discharge atmospheric pressure chemical ionization: evaluation and application to the analysis of animal drug residues in complex matrices

Alternative ionization methods are increasingly being utilized to increase the versatility and selectivity of liquid chromatography/mass spectrometry (LC/MS). One such technique is the practice of using commercially available atmospheric pressure chemical ionization (APCI) sources with the corona discharge turned off, a process termed no-discharge APCI (ND-APCI). The relative LC/MS responses for several different classes of veterinary drugs were obtained by using ND-APCI, electrospray ionization (ESI), and APCI. While the ND-APCI-MS and -MSn spectra for these compounds were comparable with ESI, ND-APCI provided advantages in sensitivity and selectivity for some compounds. Drugs that were charged in solution as cations or sodium adducts responded particularly well with this technique. Instrumental parameters such as temperatures, gas and liquid flow rates, and source design were investigated to determine their effect on the process of ND-APCI. This paper explores advantages of using ND-APCI for the determination and confirmation of drug residues that might be found in food matrices, including malachite green residues in fish tissue and avermectin residues in milk.     

A reproducing kernel condition for indeterminacy in the multidimensional moment problem

Using the smallest eigenvalues of Hankel forms associated with a multidimensional moment problem, we establish a condition equivalent to the existence of a reproducing kernel. This result is a multivariate analogue of Berg, Chen, and Ismail's 2002 result. We also present a class of measures for which the existence of a reproducing kernel implies indeterminacy.

     

Liquid chromatographic determination of malachite green and leucomalachite green (LMG) residues in salmon with in situ LMG oxidation

A liquid chromatography (LC) method is presented for the quantitative determination of malachite green (MG) in salmon. MG and leucomalachite green (LMG) residues were extracted from salmon tissue with ammonium acetate buffer and acetonitrile, and then isolated by partitioning into dichloromethane. LMG was quantitatively oxidized to the chromic MG by reaction with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone. Samples were then cleaned up by solid-phase extraction with alumina and propylsulfonic acid phases. Extracts were analyzed for MG by LC with visible detection at 618 nm using isocratic elution and a C18 column. The method was validated in 35 farm-raised salmon (Salmo salar) tissues fortified at 1, 2, 4, and 10 ng/g (ppb) with an average recovery of 95.4% and a relative standard deviation of +/- 11.1%, and in 5 canned salmon (Oncorhynchus gorbuscha) samples fortified at 10 ng/g with an average recovery of 88.9 +/- 2.6%. This study also included the determination of MG and LMG residues in tissues from salmon that had been treated with MG MG was quantitatively determined at the method detection limit of 1 ng/g.