Abstract
The inhibitory effect of new chemical entities on rat liver P450 marker activities was investigated in a functional approach towards drug development. Treatment of colorectal cancer (CRC) and chemoprevention using salicylic acid has gained a lot of attention, mainly in the prevention of the onset of colon cancer. Thus, an in vitro inhibitory effect of salicylic acid on rat CYP2C11 activity was examined by using high performance liquid chromatography (HPLC). High performance liquid chromatography analysis of a CYP2C11 assay was developed on a reversed phase C18 column (SUPELCO 25 cm × 4.6 mm × 5 µm) at 243 nm using 32% phosphate buffer (pH 3.36) and 68% methanol as a mobile phase. The CYP2C11 assay showed good linearity for all components (R2 > 0.999). Substrates and metabolites were found to be stable for up to 72 h. Additionally, the method demonstrated good reproducibility, intra- and inter-day precision (<15%), acceptable recovery and accuracy (80%–120%), and low detection (1.3501 µM and 3.2757 µM) and quantitation limit values (4.914 µM and 9.927 µM) for 16α-hydroxytestosterone and testosterone, respectively. Salicylic acid acts reversibly as a noncompetitive (weak) inhibitor with Ki = 84.582 ± 2.67 µM (concentration of inhibitor to cause 50% inhibition of original enzyme activity (IC50) = 82.70 ± 2.67 µM) for CYP2C11 enzyme activity. This indicates a low potential to cause toxicity and drug–drug interactions.
Highlights
Cytochrome P450s are known as Phase 1 mono-oxygenase drug-mediated enzymes that play a vital role as a body defence in the drug pharmacokinetic field [1]
UV-VIS spectrophotometry analysis for CYP2C11 assay was performed by dissolving salicylic acid (100 μM), testosterone (200 μM) and 16-α hydroxytestosterone (50 μM) powder in pure methanol
Potassium phosphate monobasic, potassium phosphate dibasic, phenacetin with purity greater than 98%, phosphoric acid (85% w/w), glucose-6-phosphate (G-6-P), glucose-6-phosphate dehydrogenase (G-6-PDH), EDTA (Ethylenediamine tetraacetic acid), NADP+ (Nicotinamide Adenine Dinucleotide Phosphate), magnesium chloride (MgCl2 ), testosterone, microsomes from liver pooled from male rats (Sprague-Dawley) (Gillingham, UK) and 16-alfa hydroxytestosterone were purchased from Sigma Aldrich, Co (Old Brickyard, Gillingham, UK)
Summary
Cytochrome P450s are known as Phase 1 mono-oxygenase drug-mediated enzymes that play a vital role as a body defence in the drug pharmacokinetic field [1]. Statistical results have shown that about 70%–80% of most marketed drugs are metabolised by five major Phase 1 drug mediated enzymes [4]. Induction or inhibition of CYP450 enzymes can result in either drug–drug or drug–herb interactions, and their effectiveness can be altered [5]. This leads to a therapeutic failure in the treatment [3].
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