UPLC-PDA-ESI-QTOF-MS/MS fingerprint of purified flavonoid enriched fraction of Bryophyllum pinnatum; antioxidant properties, anticholinesterase activity and in silico studies

Figures & data

Figure 1. Bryophyllum pinnatum plant.

Figure 2. Overlay of UPLC chromatogram at 280 nm top and base peak intensity (BPI) chromatograms (bottom) of BPFRF.

Figure 3. Photodiode array detector (PDA) spectra and mass spectra (MS/MS) fragmentation pattern present in B. pinnatum flavonoid-rich fraction (BPFRF). Luteolin C-glucoside-C-arabinoside (carlinoside) (A). Quercetin (B). Luteolin (C). Quercetin-3-methyl ether (isorhamnetin) (D). Luteolin-7-glucoside (E).

Table 1. Constituents of BPFRF identified and characterized by UPLC-PDA-Q/TOF-MS² analysis.

Peaks	Retention time (min)	UPLC PDA λmax (nm)	^am/z	^bM-H^−	^cMS^E(MS/MS)	Concentration g/100 g	Tentative Identification
1a	18.56	221, 348	579.1341	C26H27O15	151, 178,243, 255,271, 300,315,447	0.27	8-Arabinopyranosyl-6-glucopyranosyl luteolin (carlinoside)
2	22.54	255, 370	301.0327	C15H9O7	107,151,178,190,245,273	13.34	Quercetin
3	25.50	263, 367	285.0374	C15H9O6	107,121,151,178,187,229,257	2.01	Luteolin
4	26.03	254,370	315.0477	C16H11O7	107,148,151,164,178,227,255,277	3.49	Quercetin methyl ester (isorhamnetin)
5	26.33	–	327.2107	C18H31O5	127,151,171,183,211,229,239,291, 300, 301,315	-	Unknown
6	27.24	221,367	329.1502	C18H33O5	127,151,171,183,211,229,233,251,300, 301	-	Unknown
7	28.32	222.41	447.2006	C24H31O8	89, 151,202, 216, 241, 285, 301, 311, 351, 379	0.92	luteolin-7-glucoside
8	29.74	223.49	795.4507	C42H67O14	119,183,236,301,339, 457, 507, 615, 647, 691, 765	-	Unknown
9	31.07	No UV	633.3901	C36H57O9	85, 113, 116,151, 183, 220, 301, 339, 407, 463, 485, 559	-	Unknown
10	32.45	No UV	293.2134	C18H29O6	98, 116, 119, 121, 169, 171, 183, 235, 236,275	-	Unknown
11	33.11	No UV	617.4011	C36H57O8	85, 113, 151, 235, 253, 277, 339, 367,439, 497, 513, 579	-	Unknown
12	34.15	226	265.1402	C15H21O4	98, 116, 151, 183, 220, 255, 279	-	Unknown
13	35.81		271.2201	C13H27O8	130, 150, 171, 183, 197,221, 220, 226, 255, 235	-	Unknown
14	37.05	226	271.2214	C13H27O8	96, 98, 100, 119, 139, 183, 199, 255, 265, 293, 325, 353, 397, 465	-	Unknown
15	37.05	226	325.1812	C14H29O8	79, 96, 119, 170, 183, 184, 197, 227, 265,293	-	Unknown

^aAccurate mass detection; ^bnegative ions; ^cmass fragmentation patterns with intensity normalised to 100 for the highest fragment. Rt: Retention time; λmax: maxima absorbance.

Figure 4. 2,2-Diphenyl-1-picrylhydrazyl free radical scavenging effect of BPFRF in comparison with ascorbic acid (3.125–100 μg/mL).

Table 2. DPPH radical scavenging potential, lipid peroxidation inhibitory activity and cholinesterase inhibitory activity of BPFRF, quercetin and standards.

Samples	DPPH radical IC50 (µg/mL)	Lipid peroxidation IC50 (µg/mL)	AChE IC50 (µg/mL)	BuChE IC50 (µg/mL)
BPFRF	7.382 ± 0.79	7.182 ± 0.60	22.283 ± 0.27	33.437 ± 1.46
Ascorbic acid	10.186 ± 0.14	14.393 ± 0.54
Quercetin	13.803 ± 0.34	25.242 ± 1.97	15.528 ± 0.08	24.326 ± 0.40
Rivastigamine			7.994 ± 0.12	20.120 ± 0.72

Figure 5. Fe²⁺-induced lipid peroxidation inhibitory activity of BPFRF and the ascorbic acid.

Figure 6. Cholinesterase inhibitory activity of BPFRF AChE (A). BuChE (B).

Table 3. Binding energies of BPFRF ligands and known inhibitors against AChE and BuChE drug targets.

Compound name	PubChem ID	Chemical formula	Binding affinity (kcal/mol) AChE	Binding affinity (kcal/mol) BuChE
Luteolin-7-glucoside*	5280637	C15H10O6	−14.9	−9.8
Luteolin*	5280445	C15H10O6	−12.3	−7.4
Quercetin*	5280343	C16H12O8	−11.2	−7.8
Isorhamnetin*	5281654	C9H8O3	−10.5	−7.4
Carlinoside*	442584	C9H10O5	−8.9	−12.7
Rivastigmine^#	77991	C28H48O	−10.4	−6.9

^#Known inhibitors; *Putative BPFRF inhibitors.

Figure 7. Illustrations of molecular interactions (left: 3D and right: 2D) between the/highest binding energies of BPFRF constituents and standard against AChE target. Luteolin-7-glucoside (red) (Ai, Aii). Quercetin (orange) (Bi, Bii). Rivastigmine (purple) (Ci, Cii).

Figure 8. Illustrations of molecular interactions (left: 3D and right: 2D) between the highest binding energies of BPFRF constituents and standard against BuChE target. Carlinoside (yellow) (Ai, Aii). Quercetin (grey) (Bi, Bii). Rivastigmine (purple) (Ci, Cii).

Data availability statement

The data that support the findings of this study are available from the authors upon request.