Anticarcinogenic activity of Muntingia calabura leaves methanol extract against the azoxymethane-induced colon cancer in rats involved modulation of the colonic antioxidant system partly by flavonoids

Figures & data

Table

Groups		Administration
Control	I (Normal control)	Normal saline + normal saline
	II (AOM control)	AOM + 8% Tween-80
Treatment	III	AOM + 50 mg/kg MEMC
	IV	AOM + 250 mg/kg MEMC
	V	AOM + 500 mg/kg MEMC

Figure 1. Body weight for male Sprague–Dawley rats for colon cancer chemopreventive study.

Figure 2. (a) Normal histoarchitecture of rat colon; (b) AOM control group; (c) section of colon tissue treated with AOM + 50 mg/kg MEMC; (d) section of colon tissue treated with AOM + 250 mg/kg MEMC; (e) section of colon tissue treated with AOM + 500 mg/kg MEMC (I – Infiltration of inflammatory cells. Black arrow – Aberrant crypt foci) (100 × magnification).

Table 1. Effect of MEMC on relative organ weigh (ROW) in AOM-treated rats.

% Organ weight/body weight	Normal control (normal saline alone)	Positive control (AOM alone)	AOM + 50mg/kg MEMC	AOM + 250mg/kg MEMC	AOM + 500 mg/kg MEMC
Spleen	0.232 ± 0.0398	0.257 ± 0.0296	0.275 ± 0.0161	0.295 ± 0.0361	0.347 ± 0.0186*
Heart	0.434 ± 0.0376	0.409 ± 0.0204	0.425 ± 0.0189	0356 ± 0.0224	0.434 ± 0.0328
Liver	2.654 ± 0.2637	2.951 ± 0.2124	3.037 ± 0.1840	2.868 ± 0.2441	3.520 ± 0.3584
Kidney	0.7814 ± 0.0552	0.796 ± 0.0295	0.762 ± 0.0251	0.804 ± 0.0334	0.982 ± 0.0676*
Lung	0.823 ± 0.0764	0.951 ± 0.0689	0.826 ± 0.0634	0.940 ± 0.1161	1.141 ± 0.1247
Colon	0.809 ± 0.0659	0.934 ± 0.0719	0.859 ± 0.0674	0.856 ± 0.0893	1.048 ± 0.1128

Values are expressed as means ± SEM.

* Significant different as compared to normal control, p < 0.05.

Table 2. The incidence of aberrant crypt foci (ACF) found in rats colon induced by azoxymethane (AOM).

				Incidence of ACF		Reduction as compared
Group	Treatment	Proximal	Middle	Distal	Total of ACF	to AOM control (%)
I	Normal control	0.00 ± 0.00	0.00 ± 0.00	0.00 ± 0.00	0.00 ± 0.00
II	AOM control	33.00 ± 3.055	27.0 0 ± 2.887	47.67 ± 3.480	107.7 ± 6.489
III	AOM + 50 mg/kg MEMC	24.67 ± 1.333*	27.33 ± 4.910	33.33 ± 5.696	85.33 ± 9.563	20.77%
IV	AOM + 250 mg/kg MEMC	23.0 ± 0.577**	25.0 ± 2.517	28.0 ± 3.512*	76.0 ± 6.000	29.43%
V	AOM + 500 mg/kg MEMC	14.33 ± 1.453***	19.67 ± 1.667	23.00 ± 4.509**	48.33 ± 13.48**	55.13%

Values are expressed as means ± S.E.M.

* Significant different as compared to AOM control, p < 0.05.

** Significant different as compared to AOM control, p < 0.01.

*** Significant different as compared to AOM control, p < 0.001.

Table 3. Effect of antioxidant enzymes in the colons of rats subjected to MEMC treatment.

			Antioxidant assays
Group	Treatment	SOD (U/mg protein)	CAT (nmol/min/mL)	GSH (µM/mg protein)	MDA (ng/g tissue)
AOM control	AOM	0.6473 ± 0.2107	95.33 ± 1.202	16.55 ± 1.913	3.097 ± 0.2621
Normal control	Normal saline	2.672 ± 0.1521***	103.4 ± 0.1764	39.57 ± 4.534***	1.520 ± 0.2403**
Treatment	AOM + 50 mg/kg MEMC	2.104 ± 0.0610***	101.9 ± 0.03333	23.13 ± 0.777*	2.460 ± 0.2879*
Treatment	AOM + 250 mg/kg MEMC	2.352 ± 0.0706***	102.7 ± 0.0*	25.41 ± 1.150*	2.343 ± 0.2418*
Treatment	AOM + 500 mg/kg MEMC	2.526 ± 0.0878***	108.1 ± 4.942**	32.06 ± 1.782**	1.200 ± 0.1015***

Values are expressed as means ± S.E.M.

* Significant different as compared to AOM control, p < 0.05.

** Significant different as compared to AOM control, p < 0.01.

*** Significant different as compared to AOM control, p < 0.001.

Figure 4. Total ion chromatography (TIC) of MEMC obtained using the UHPLC instrument in negative ion mode.

Table 4. Phenolic compounds identified in MMCL by UHPLC-MS.

Peak no.	t^R (min)	[M–H]–	Error (ppm)	Formula	Identification
1.	0.46	169.01392	5.090	C7H5O5	Gallic acid
2.	3.16	193.05023	3.599	C10H9O4	Ferulic acid
3.	4.58	599.10559	4.079	C28H23O15	Quercitrin-2″-O-gallate
4.	5.08	447.09439	4.926	C21H19O11	Kaempferol-3-O-galactoside
5.	5.39	463.08957	5.392	C15H9O8	Quercetin-3-O-galactoside
6.	6.96	583.11093	4.576	C28H23O14	Afzelin-O-gallate
7.	7.44	603.07947	4.209	C30H19O14	Quercetin dimer
8.	8.12	593.13110	3.596	C30H25O13	Kaempferol-3-O-glucoside
9.	8.57	271.06100	3.321	C15H12O5	Pinobaksin
10.	11.45	255.06644	4.919	C15H11O4	Pinocembrin (isomer 2)
11.	11.92	299.05606	3.496	C16H11O6	Kaempferide I
12.	12.39	299.05649	4.934	C16H11O6	Kaempferide II
13.	12.78	313.07214	4.713	C17H13O6	Ermanin II