Differential Induction of Flavonoids in Groundnut in Response to Helicoverpa armigera and Aphis craccivora Infestation

Flavonoids are important plant secondary metabolites, which protect plants from various stresses, including herbivory. Plants differentially respond to insects with different modes of action. High performance liquid chromatography (HPLC) fingerprinting of phenols of groundnut ( Arachis hypogaea) plants with differential levels of resistance was carried out in response to Helicoverpa armigera (chewing insect) and Aphis craccivora (sucking pest) infestation. The genotypes used were ICGV 86699, ICGV 86031, ICG 2271 (NCAc 343), ICG 1697 (NCAc 17090), and JL 24. Most of the identified compounds were present in H. armigera- and A. craccivora-infested plants of ICGV 86699. Syringic acid was observed in all the genotypes across the treatments, except in the uninfested control plants of ICG 2271 and aphid-infested plants of ICG 1697. Caffeic acid and umbelliferone were observed only in the H. armigera-infested plants of ICGV 86699. Similarly, dihydroxybenzoic acid and vanillic acid were observed in H. armigera- and aphid-infested plants of ICG 2271 and JL 24, respectively. The peak areas were transformed into the amounts of compounds by using internal standard peak areas and were expressed in nanograms. Quantities of the identified compounds varied across genotypes and treatments. The common compounds observed were chlorogenic, syringic, quercetin, and ferulic acids. These results suggest that depending on the mode of feeding, flavonoids are induced differentially in groundnut plants.

Liquid-chromatographic determination of dolichols in urine.

A reversed-phase "high-performance" liquid chromatographic assay for dolichols-18, -19 and -20 in urine is described. Dolichols are extracted from urine by using C18 cartridges and are chromatographed with a mobile phase consisting of 2-propanol/methanol, the effluent being monitored at 210 nm. The useful lower limit of sensitivity for quantification is 4 pmol (5 ng) of each dolichol per 5-microL injection, corresponding to 1.6 nmol (2 ng) per liter of urine. Heneicosaprenol is satisfactory as the internal standard. Peak heights and the amounts of dolichols applied to the column are linearly related from 4 to 110 pmol. Mean analytical recovery was 71%. For three different concentrations the mean within-assay CV was 6.4%, the between-assay CV 11%. The normal reference interval of total dolichols found for healthy adults was 17-101 micrograms/24 h (n = 30) or 1.9-11 micrograms per millimole of creatinine (n = 39). I determined the distribution of the main dolichols in urine and applied the assay also for samples from alcoholics.

Liquid-chromatographic determination of total hydroxyproline in urine.

A reversed-phase "high-performance" liquid chromatographic assay for total hydroxyproline in urine is described. The urine samples are hydrolyzed overnight with acid, evaporated, solubilized, and derivatized with 4-dimethylaminoazobenzene-4'-sulfonyl chloride. The derivatives are chromatographed with a solvent gradient consisting of sodium acetate buffer and acetonitrile, the effluent being monitored at 436 nm. The useful lower limit of sensitivity for quantification is 13 pmol of hydroxyproline per 5-microL injection, corresponding to 33 mumol/L of urine. Either glutamine or cysteic acid is satisfactory as the internal standard. Peak heights and the amounts of hydroxyproline applied to the column are linearly related from 13.3 to 266 pmol. Mean analytical recovery was 83%. For four different concentrations the mean within-assay CV was 9.0% and the between-assay CV 12%. The normal reference interval found for 31 healthy adults was 31 to 177 mumol/24 h per square meter of body surface. We compared results for 10 samples from patients.

Improved liquid-chromatographic determination of mexiletine, an antiarrhythmic drug, in plasma.

In this improved reversed-phase liquid-chromatographic procedure for determination of mexiletine in plasma, mexiletine and an internal standard, chlorodisopyramide, are extracted with methylene chloride from 0.5 mL of serum or plasma; the extract is then concentrated and injected onto a C18 chromatographic column. Mexiletine in the column effluent is detected by monitoring absorbance at 210 nm. It is quantified by use of mexiletine-internal standard peak-height ratios. The relation between this ratio and mexiletine concentration is linear from 0.1 to 5.0 mg/L. The lower limit of detection is about 50 micrograms/L. At a mexiletine concentration of 2.0 mg/L in serum, intrarun precision (CV) is 2.9% and inter-run precision is 5.9%; at 0.5 mg/L, these CVs are 5.7% and 9.6%, respectively. Analytical recovery of added mexiletine in serum is 68-88%. Therapeutic concentrations of some commonly administered drugs in patients' specimens did not interfere. In serum from 38 patients receiving mexiletine for cardiac arrhythmia, concentrations measured by this method correlated with therapeutic efficacy.

Determination of Docosenoic Acids in Fats and Oils by Gas-Liquid Chromatography

A rapid gas-liquid chromatographic (GLC) procedure has been developed for the determination of docosenoic acids in fats and oils. The method involves conversion of a known weight of oil to methyl esters using sodium methoxide-methanol, with a known weight of methyl tetracosanoate used as an internal standard. After acidification and extraction into hexane, esters are analyzed by GLC on a diethylene glycol succinate column. The percentage of docosenoic acids is calculated from the docosenoate and internal standard peak areas. The method is particularly suited to the determination of low levels (<5%) of docosenoic acids.

Gas-Liquid Chromatographic Determination of Diethylstilbestrol Monomethyl Ether in Diethylstilbestrol

A gas chromatographic method is described for the determination of the monomethyl ether of diethylstilbestrol (MME) in diethylstilbestrol (DES). A 1 g sample of DES is dissolved in an aqueous solution of 0.5N sodium hydroxide and extracted with a chloroform solution containing benefin, the internal standard. The chloroform solution is back-extracted with sodium hydroxide, and an aliquot is chromatographed after silylation. The MME peaks (cis and trans') are measured relative to the internal standard peak. The method was tested for accuracy within the 120—2000 ppm range, and a linear plot of detector response vs. concentration was obtained. The method is also suitable for the determination of diethylstilbestrol dimethyl ether (DME) if present with MME in a DES sample.

Gas Chromatographic Determination of Diethylstilbestrol Dimethyl Ether in Diethylstilbestrol

A GLC method is described for the determination of the dimethyl ether of diethylstilbestrol (DME) in diethylstilbestrol (DES). A 1 g sample of DES is dissolved in an aqueous solution of 0.5N NaOH and then extracted with a chloroform solution containing benefin, the internal standard. An aliquot of the chloroform layer is chromatographed and the DME peak is measured relative to the internal standard peak. The method was tested for accuracy within the 11–110 ppm range and a linear plot of detector response vs. concentration was obtained.