Carbonyl Reduction of Mequindox by Chicken and Porcine Cytosol and Cloned Carbonyl Reductase 1

Abstract

ABSTRACT:Mequindox (MEQ) is a novel synthetic quinoxaline 1,4-dioxides derivative, which is widely used as a veterinary drug and animal feed additive. However, the metabolic mechanism of MEQ is rarely reported. The N-oxide reduction mechanism of MEQ was reported in our previous work. In this article, the toxicity and the reduction of the carbonyl of MEQ were studied. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium assays demonstrated that the carbonyl-reduced MEQ, 2-isoethanol MEQ was much less toxic than MEQ. High-performance liquid chromatography analysis showed that the cytosol extracts of chicken and pig livers were able to reduce MEQ to 2-isoethanol MEQ and the reaction was NADPH-dependent. Further study via enzymeinhibitory experiment revealed that carbonyl reductase 1 (CBR1) participated in this metabolism. The enzyme activity analysis showed that both chicken CBR1 (cCBR1) and porcine CBR1 (pCBR1) were capable of catalyzing the carbonyl reduction of MEQ and its N-oxide reductive metabolite, 1-deoxymequindox. By comparison of the kinetic constants, we observed that the activity of cCBR1 was higher than pCBR1 to MEQ and the standard substrate of CBR1, menadione. On the other hand, both CBR1s exhibited higher activity to 1-deoxymequindox than MEQ. Mutation analysis suggested that the difference of amino acid at position 141/142 may be one possible reason that caused the activity difference between cCBR1 and pCBR1. Thus far, CBR1 was first reported to participate in the carbonyl reduction of MEQ. Our results will be helpful to recognize the metabolic pathways of quinoxaline drugs deeply and to provide a theoretical basis for controlling the negative effects of these drugs.

The critical role of oxidative stress in the toxicity and metabolism of quinoxaline 1,4-di-N-oxides in vitro and in vivo

“…AO, acridine orange; cCBR1, chicken carbonyl reductase 1; porcine carbonyl reductase 1; SsAOX1, porcine aldehyde oxidase. Cited from (Chen et al, 2013;Huang et al, 2015bHuang et al, , 2015cLi et al, 2014aLi et al, , 2014bLiu et al, 2008Liu et al, , 2009Liu et al, , 2010aLiu et al, , 2010bLiu et al, , 2010cLiu et al, , 2011aLiu et al, , 2011bLiu & Sun, 2013;Mu et al, 2014;Tang et al, 2012;Wang et al, 2016b;Yang et al, 2013a;Zheng et al, 2011). marked as less genotoxicity than other kinds Wang et al, , 2011b.…”

“…Carbonyl reductase 1 (CBR1) was reported to participate in the carbonyl reduction of MEQ, and CBR1 was able to reduce MEQ to 2-isoethanol MEQ and the reaction was NADPH-dependent. Both chicken CBR1 (cCBR1) and porcine CBR1 (pCBR1) were capable of catalyzing the carbonyl reduction of MEQ and its N-oxide reductive metabolite, 1-deoxy MEQ (Tang et al, 2012). Furthermore, the activity of cCBR1 was higher than pCBR1 to MEQ, which might be due to the different amino acids at position 141/142 (Tang et al, 2012).…”

“…Both chicken CBR1 (cCBR1) and porcine CBR1 (pCBR1) were capable of catalyzing the carbonyl reduction of MEQ and its N-oxide reductive metabolite, 1-deoxy MEQ (Tang et al, 2012). Furthermore, the activity of cCBR1 was higher than pCBR1 to MEQ, which might be due to the different amino acids at position 141/142 (Tang et al, 2012). In chicken (Gallus gallus), both CYP1A4 and CYP1A5 participate in the oxidative metabolism of QdNOs.…”

“…The N!O reduction metabolite of MEQ, 2-isoethanol 4-desoxymequindox, was detected in the liver and spleen with an increase of SOD, and in the testes following the appearance of 8-OhdG after rats were exposed to MEQ (25, 55, 110 and 275 mg/kg diet) for 180 days, suggesting that 2-isoethanol 4-desoxymequindox might be involved in the organ toxicity induced by MEQ (Ihsan et al, 2011;Wang et al, 2011c). The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) assays demonstrated that the carbonyl-reduced MEQ, 2-isoethanol MEQ was much less toxic than MEQ (Tang et al, 2012). However, desoxycarbadox caused chromosome damage with 10 mg/kg b.w.…”

Dynamic changes in metabolic profiles of rats subchronically exposed to mequindox

“…1), a synthetic quinoxaline 1,4-dioxides (QdNOs) derivative, was developed as a broad-spectrum antibacterial veterinary drug 1,2 and is widely used as an animal feeding additive in China. 3,4 The pharmacokinetics of the drug have been studied in a variety of species, e.g. rabbit, 5 sheep, 6 goat, 7 swine, 8 chicken 1 and rat.…”