Synthesis of chiral pyrazolo[4,3-e][1,2,4]triazine sulfonamides with tyrosinase and urease inhibitory activity

Figures & data

Scheme 1. Synthetic pathway to the sulfonamides 8a–i. Reagents and conditions: (a) CH₃CH₂NO₂, KOH, DMSO, 2 h, 80–86%; (b) Na₂S₂O₄, H₂O/dioxane, rt, 12 h, 55–65%; (c) CH₃NH–NH₂, PTSA, EtOH, rt, 1 h, 50–55%; (d) method A: 10% HCl, EtOH, reflux, 1 h, 58–61%; method B: PTSA, 140 °C, 1 min, 61%; (e) ethoxyphenylboronic acid, Pd(PPh₃)₄, CuMeSal, THF, Ar, reflux, overnight, 75–80%; (f); ClSO₃H, 0 °C to rt, 2 h, 75–95%; (g) appropriate amine, anhydrous MeCN, rt, overnight, 72–93%.

Table 1. The inhibitory effects of compounds 8a–8j on mushroom tyrosinase and urease.

Compounds code	Mushroom tyrosinase IC50 (μM)	Urease IC50 (μM)
8a	38.46 ± 0.42	0.037±0.003
8b	30.76±3.56	0.084 ± 0.020
8c	39.00 ± 1.34	0.078 ± 0.020
8d	38.15 ± 1.68	0.064 ± 0.020
8e	39.75 ± 1.92	0.076 ± 0.007
8f	34.72 ± 2.01	0.044±0.130
8g	36.49 ± 1.75	0.063 ± 0.016
8h	32.46 ± 1.87	0.080 ± 0.014
8i	40.17 ± 1.09	0.042±0.012
8j	27.9±2.43	0.080 ± 0.015
Kojic Acid	16.69 ± 2.8	–
Thio-Urea	–	20.7 ± 0.45

Bolded values represent the most active derivatives.

Figure 1. a) Lineweaver–Burk plots for the inhibition of mushroom tyrosinase in the presence of compound (8b). Concentrations of (8b) were 0, 15, 30, 61, 123 and 247 μM, respectively. Substrate l-DOPA Concentrations were 0.125, 0.25, 0.5, 1 and 2 mM, respectively. b) The secondary replot of the Lineweaver–Burk plot, 1/V (y-intercept) of a) versus various concentrations of (8b). c) The Dixon plot of the reciprocal of the initial velocities versus various concentrations of compound (8b) at fixed substrate concentration.

Figure 2. a) Lineweaver–Burk plots for the inhibition of mushroom tyrosinase in the presence of compound (8j). Concentrations of (8j) were 0, 14, 28 and 55.8 μM, respectively. Substrate l-DOPA Concentrations were 0.062, 0.125, 0.25, 0.5, 1 and 2 mM, respectively. b) The Dixon plot of the reciprocal of initial velocities versus various concentrations of compound (8j) at fixed substrate concentration.

Figure 3. a) Double reciprocal Lineweaver–Burk plots for the inhibition of Jack bean urease in the presence of compound (8a). Concentrations of (8a) were 0, 0.125, 0.25 and 0.5 μM, respectively. Substrate urea concentrations were 3.12, 6.25, 12.5, 25, 50 and 100 mM, respectively. b) The secondary replot of the Lineweaver–Burk plot, 1/V (y-intercept) of a) versus various concentrations of (8a). c) The Dixon plot of the reciprocal of the initial velocities versus various concentrations of compound (8a) at fixed substrate concentration.

Table 2. Inhibitory effect of compound 8b and 8j on mushroom tyrosinase activity and of 8a on urease activity.

Compound	Enzyme
	Mushroom tyrosinase		Urease
	Ki (μM)	Type of inhibition	Ki (μM)	Type of inhibition
	8a	–	–	0.01	Mixed type
8b	40	Noncompetitive	–	–
8j	20	Mixed type	–	–

Ki is enzyme inhibition constant.

– is not determined.