Accurate Analysis of Tricarboxylic Acid Cycle Metabolites and Anion Components in Hemocytes by IC-CD/ESI-MS for Quantifying Insecticide Impairment on Cellular Immunity in Mythimna separata

Abstract

Insecticides are more broadly known to affect insect cellular immunity, but the components in hemocytes and their response to insecticide stress are still unknown. In this paper, a method based on trifluoroacetic acid extraction, followed by IC-CD/ESI-MS analysis, was developed to simultaneously determine tricarboxylic acid (TCA) cycle metabolites and anion components in hemocytes from Mythimna separata larvae. Validation gave excellent selectivity, recovery (88.7–107.6%), linear correlation (r 2 > 0.9961), precision (<3.89%), LOD (0.002–0.006 mg/L), LOQ (0.006–0.020 mg/L), and a short chromatographic run. The method was verified by administration of 4-((3-chloro-4-fluorophenyl)­amino)-7-methoxyquinazolin-6-yl 3-(1,3-dioxoiso-indolin-2-yl) propanoate (QDP) or emamectin benzoate (EMB) to hemocytes in vitro and larvae in vivo. TCA metabolites including citrate, α-ketoglutarate, fumarate, malate, and oxaloacetate, and anions including acetate, oxalate, chloride, carbonate, and sulfate were identified and clearly separated. QDP and EMB showed a biphasic dose effect on TCA metabolites, and the contrary hormesis paralleled the different actions of QDP and EMB. The inhibition or improvement of cellular immunity depended on the QDP concentration. In conclusion, a highly sensitive, reliable, and robust method was developed, enabling the monitoring of hemocyte immunity by the quantification of TCA metabolites and anion components in minute hemocyte samples.

Real-Time Monitoring of Translocation, Dissipation, and Cumulative Risk of Maleic Hydrazide in Potato Plants and Tubers by Ion Exclusion Chromatography

“…For example, MH residues were determined in cured tobacco by gas chromatography, in garlic bulbs, potatoes, and onions by high-performance liquid chromatography, , and in tobacco and grapes by liquid chromatography with tandem mass spectrometry. , The improvement in instrumentation and analytical techniques has provided more sensitive detection of MH residues. , However, MH translocation and distribution in plants have been less studied due to the technical difficulties associated with simultaneous multiple tissues and soil experiments . Ion chromatography (IC) is a powerful analytical tool in the detection of small-molecule organic compounds and enables the multiresidue simultaneous determination of polar anionic pesticides in fruit and vegetables. , The IC techniques offer an enormous range of possibilities for selecting stationary and mobile phases and have a broad range of application in agriculture, pharmaceutical, and clinical studies because of ease of operation, high sensitivity, and online performance. , We previously reported the use of an effective IC method for real-time accurate analysis of tricarboxylic acid cycle metabolites in insect hemocytes . Herein, we will develop a novel method using the IC techniques that are suitable for the determination of MH in a variety of samples such as leaves, stems, roots, tubers, and buds of potatoes and rhizosphere soil to realize real-time monitoring of MH translocation behavior.…”

“…27,28 We previously reported the use of an effective IC method for realtime accurate analysis of tricarboxylic acid cycle metabolites in insect hemocytes. 29 Herein, we will develop a novel method The importance of understanding MH translocation behavior is to enable us to predict its time-dependent level remaining in different plant tissues. In this paper, we aimed to provide a real-time insight into MH translocation behavior in potato plants, roots, and tubers and examine the dynamics of MH accumulation and dissipation in plant tissues.…”