Major chemical constituents and antioxidant activities of different extracts from the peduncles of Hovenia acerba Lindl

Figures & data

Table 1. The total phenolic, tannin contents, and antioxidant activities of HAP extracted by different solvents^a.

Samples	Total phenolic content (mg of GAE/g DW)^b	Total tannin content (mg of CAE/g DW)^c	DPPH assay (μmol of TE/g DW)^d	ABTS assay (μmol of TE/g DW)^d	FRAP assay (μmol of FE/g DW)^e
Water extract	5.06 ± 0.10^b	6.78 ± 0.68^b	31.25 ± 1.88^b	22.06 ± 2.12^b	20.47 ± 0.99^b
80% methanol extract	12.46 ± 0.60^a	11.17 ± 0.53^a	60.95 ± 0.12^a	59.38 ± 2.88^a	82.43 ± 2.97^a
Ethyl acetate extract	1.12 ± 0.04^c	1.03 ± 0.03^c	2.10 ± 0.02^d	1.21 ± 0.03^c	4.72 ± 0.36^c
Hexane extract	1.24 ± 0.03^c	5.46 ± 0.24^b	5.31 ± 0.43^c	3.62 ± 0.23^c	4.32 ± 0.14^c

^aResults were expressed as mean ± standard deviation of three replicates. Values followed by the different letters (a, b, c, d) within the same column are significantly different (P < 0.05).

^bPhenolic contents were expressed as mg of gallic acid equivalents (GAE) in 1 g of dry weight (DW) of plant material ± standard deviation.

^cTannin contents were expressed as mg of catechin equivalents (CAE) in 1 g of dry weight of plant material ± standard deviation.

^dDPPH and ABTS radical-scavenging activities of the sample were compared with a Trolox standard curve, and results were expressed as μmol of Trolox equivalents (TE) in 1 g of dry weight material ± standard deviation.

^eFRAP assay was compared with a FeSO₄ standard curve, and results were expressed as μmol of FeSO₄ Equivalents (FE) in 1 g of dry weight material ± standard deviation.

Table 2. Characterization of the main phytochemicals in different HAP extracts by UPLC-ESI-QTOF-MS/MS.

				[M-H]^−/[M+ HCOOH-H]^− (m/z)^A
Compound no.	tR (min)	λmax (nm)	Formula	Measured	Calculated	Error (ppm)	Major fragment ions (m/z)	Identification	Distribution
Phenolic acids and derivatives
15	7.698	278	C7H6O4	153.0193	153.0193	0.21	109.0129[M-H-CO2]^−	Protocatechuic acid^a,b,c	ME, EE, WE
18	7.848	278	C13H16O9	315.0730	315.0722	0.49	153.0125[M-H-C6H10O5]^−, 109.0126[M-H-C6H10O5-CO2]^−	Protocatechuic acid hexoside^a,b	ME, EE, WE
19	8.464	310	C14H18O9	329.087	329.0878	2.44		3-(b-D-glucopyranosyloxy)-4-hydroxy-benzoic acid-methyl ester(woodorien)^a,b	ME, WE
20	8.581	280/305	C10H8O6	223.0243	223.0248	2.28		2,4-Dihydroxy-6-pyruvylbenzoic acid ^b	ME, WE
43	9.696	260/295	C16H18O9	353.0881	353.0878	−0.83	191.0576[M−H-C9H6O3]^−	3-Caffeoylquinic acid^a,b,c	ME, WE
44	10.348	255	C7H6O3	137.0240	137.0244	3.03		p-Hydroxybenzoic acid^a,b,c	ME, EE, WE
54	12.77	218/274	C9H10O5	197.0444	197.0455	5.76		Ethyl gallate^a,b	ME, WE
57	13.716	296/324	C10H10O4	193.0501	193.0506	2.74	178.0261[M-H-CH3]^−, 149.0584[M-H-CO2]^−, 134.0361[M-H-CH3-CO2]^−	Ferulic acid^a,b,c	ME, EE, WE
62	17.523	269/330	C16H18O8	337.0933	337.0929	−1.21		4-p-Coumaroylquinic acid^a,b	ME, WE
63	17.007	296/324	C10H10O4	193.0504	193.0506	1.2	178.0261[M-H-CH3]^−,	Isoferulic acid^a,b,c	ME, EE, WE
							149.0584[M-H-CO2]^−,
							134.0361[M-H-CH3-CO2]^−
110	18.695	270/340	C21H30O12	473.1673	473.1664	−1.79	341.1234[M-H-C5H8O4]^−, 179.0695[M-H-C6H10O5-C5H8O4]^−, 164.0471[M-H-C11H18O9-CH3]^−	3-(4-Methoxyphenyl)propanoic acid vicianoside^b	ME, WE
Flavonoids
Flavones and derivatives
47	11.099	270/350	C33H40O21	771.1944	771.1989	0.43	609.2587[M-H-C6H10O5]^−, 463.1135[M-H-C12H20O9]^−, 300.0258[M-H-C12H21O9-C6H10O5]^− 301.0333[M-H-C12H20O9-C6H10O5]^−	Quercetin 3-glucoside-7-rutinoside ^a,b	ME, WE
51	12.277	278	C16H12O8	331.0452	331.0459	2.23	303.0518[M-H-CO]^−	3ʹ-O-Methylmyricetin^a,b	ME
52	12.344	280	C33H40O20	755.2046	755.2040	−0.77	609.2198[M-H-C6H10O4]^−, 463.0861[M-H-C12H20O8]^−, 447.0959[M-H-C12H20O9]^−	Quercetin 3-Neohesperidoside-7-rhamnoside^a,b,c	ME, WE
53	12.66	275	C27H30O17	625.1405	625.1410	0.84		Myricetin-rutinoside^a,b	ME, WE
55	12.86	275	C21H20O13	479.0834	479.0831	−0.6		Myricetin-hexoside^a,b	ME, WE
59	14.709	270	C27H30O16	609.1459	609.1461	0.34	300.0264[M-H-C12H21O9]^−, 301.0337[M-H-C12H20O9]^−, 271.0243[M-H-C12H21O9-CHO]^−, 255.0285[M-H-C12H20O9-CO-H2O]^−, 151.0041[M-H-C12H21O9-C8H5O3]^−	Rutin ^a,b,c	ME, WE
60	14.959	280/335	C21H20O12	463.0882	463.0882	0	317.0648[M-H-C6H10O5]^−	Myricetin-rhamnoside^a,b	ME, WE
61	16.874	270/340	C15H10O8	317.0304	317.0303	−0.34		Myricetin ^a,b,c	ME
69	31.345	268/317	C15H10O4	253.0506	253.0496	4.06		Chrysin^a,b,c	EE
70	32.203	278/339	C15H10O5	269.0451	269.0455	1.66	239.0335[M-H-CH2O]^−, 227.0335[M-H-CO-CH2]^−	Apigenin^a,b,c	EE
117	30.06	281	C17H14O6	313.0717	313.0718	0.2		Kumatakenin^a,b	ME, EE
118	32.875	279	C18H16O6	327.0871	323.0874	0.95	295.0970[M-H-CH2-H2O]-	3-Hydroxy-2ʹ,4ʹ,7-trimethoxyflavone^a,b	EE
119	32.741	261	C16H12O5	283.0608	283.0612	1.4		Genistein^a,b,c	EE
124	18.922	280/325	C15H10O7	301.0354	301.0357	−1.07		Quercetin^a,b,c	ME, EE
Flavanones and derivatives
24	9.08	270/302	C27H32O16	611.1620	611.1618	−0.39	465.1036[M-H-C6H10O4]^−, 303.0498[M-H-C12H20O9]^−	Dihydroquercetin-O-rutinoside^a,b	ME
46	10.608	283	C15H12O8	319.0462	319.0459	−0.81	301.0352[M-H-H2O]^−,	Amepelopsin (Dihydromyricetin) ^a,b,c	ME, EE, WE
							193.0126[M-H-C6H6O3]^−,
							151.0039[M-H-C8H8O4]^−,
							125.0037[M-H-C6H10O7]^−
56	12.927	260/310	C15H12O7	303.0513	303.0510	−0.9	285.0290[M-H-H2O]^−,	(±)-Dihydroquercetin (taxifolin)^a,b,c	ME, WE
							151.0020[M-H-C8H8O3]^−,
							125.0232[M-H-C9H6O4]^−
115	22.519	280	C15H12O5	271.0606	271.0612	2.19		Naringenin^a,b,c	EE
116	29.994	274	C15H12O4	255.0664	255.0663	−0.46		Dihydrochrysin^a,b	ME, EE
123	7.282	270/330	C21H22O10	433.1141	433.1140	0.05		Prunin(naringenin-7-O-glucoside)^a,b,c	ME, EE, WE
Flavanes and derivatives
11	6.016	280	C30H26O14	609.1245	609.1250	0.79	305.0666[M-H-C15H12O7]^−	Epigallocatechin dimer^a,b	ME, WE
13	7.132	280	C30H26O13	593.1288	593.1301	2.13	305.0666[M-H-C15H12O6]^−	Epigallocatechin derivative	ME, WE
								((2R,3S)-8-[(2R,3S,4R)-3,7-dihydroxy-2-
								(3,4,5-trihydroxyphenyl)-3,4-dihydro-2H-
								Chromen-4-yl]-2-(3,4,5-trihydroxyphenyl)-
								3,4-dihydro-2H-chromene-3,5,7-triol)^a,b
17	7.815	278	C30H26O12	577.1358	577.1351	−1.12	289.0720[M-H-C15H12O6]^−	Procyanidin B^a,b	ME, EE, WE
21	8.73	282	C15H14O7	305.0664	305.0667	0.9	287.0540 [M-H-H2O]^−,	Epigallocatechin^a,b,c	ME, WE
							261.0733 [M-H-CO2]^−,
							219.0652[M-H-C3H2O3]^−,
							179.0325[M-H-C6H6O3]^−,
							167.0331[M-H-C7H6O3]^−,
							137.0228[M-H-C8H8O4]^−,
							125.0234[M-H-C9H8O4]^−,
							111.0442[M-H-C9H6O5]^−
22	9.245	278	C15H14O6	289.0723	289.0718	−1.86	271.0615[M-H-H2O]^−,	Epicatechin^a,b,c	ME, EE, WE
							245.0833[M-H-CO2]^−,
							205.0482 [M-H-C4H4O2]^−,
							179.0350[M-H-C6H6O2]^−,
							165.0166[M-H-C7H8O2]^−
							149.0242[M-H-C7H8O3]^−
109	7.121	234/285	C15H14O7	305.0669	305.0667	−0.73		Gallocatechin^a,b,c	ME, EE, WE
122	8.997	278	C15H14O6	289.0715	289.0718	0.9		Catechin^a,b,c	ME, EE, WE
Aurones and derivatives
42	9.963	290	C27H32O15	595.1666	595.1668	0.41	577.1613[M-H-H2O]^−, 549.2634[M-H-CO-H2O]-,	Maesopsin 4-O-glucosido-4ʹ-O- rhamnoside (Hovetrichoside D)^a,b	ME
							505.1338[M-H-C4H10O2]^−,
							475.1285[M-H-C5H10O3]^−, 415.1047[M-H-C9H6O4]^−, 385.0939[M-H-C7H14O7]^−, 355.0827[M-H-C8H16O8]^−
45	10.735	298	C21H22O11	449.1082	449.1089	1.63	287.0557[M-H- C6H10O5]^−,	Maesopsin 4-O-glucoside (Hovetrichoside C)^a,b	ME, EE
							269.0432[M-H-C6H10O5-H2O]^−,
							259.0607[M-H- C6H10O5-CO]^−
64	15.275	220	C15H12O6	287.056	287.0561	0.39		2,4,6-Trihydroxy-2-(4-hydroxybenzyl)-1-benzofuran-3(2h)-one (maesopsin)^a,b	ME
Triterpenes and steroids
7	31.67	n.d	C21H34O7	397.2237	397.2232	−1.06	351.2189[M-H-HCOOH]^−, 201.1118[M-H-C11H16O3]^−, 193.0844[M-H-C10H20O4]^−, 171.1025[M-H-C12H18O4]^−	Stephanol^b	ME, EE
49	55.989	n.d.	C39H54O6	617.384	617.3848	1.23		Obtusin^b	ME, EE
72	32.344	220	C30H48O5	487.3411	487.3429	3.68		Hovenic acid^b	ME, EE, WE
73	32.477	192/220	C30H44O7	515.3002	515.3014	2.38	561.3066[M-H+ HCOOH]^−,	Ceanothetric acid^b	ME, EE, WE
75	34.675	220	C32H48O7	543.3312	543.3327	2.81		Hovenidulcigenin A^b	ME, EE
76	35.871	220	C30H48O4	471.3478	471.348	0.39	517.3439[M-H+ HCOOH]^−,	Jujubogenin^b	ME, EE
79	38.172	n.d	C30H44O5	483.3119	483.3119	−0.62	439.3198[M-H-CO2]^−, 395.3322[M-H-2CO2]^−, 325.2519[M-H-C7H10O4]^−	Fupenzic acid^b	ME, EE, HE
81	39.439	n.d.	C30H46O5	485.3276	485.3272	−0.72	971.6625[2M-H]-,	Ceanothic acid^b	ME, EE, WE
							531.5153[M+ HCOO^−]^−
83	42.619	265	C39H54O7	633.3793	633.3797	0.6		3-O-Coumaroylarjunolic acid^b	ME, EE
84	44.85	n.d.	C30H46O4	469.3319	469.3323	0.5	515.3379[M-H+ HCOOH]^−,	Gypsogenin^b	ME, EE, HE
							939.6724[2M-H]^−
86	48.23	221	C29H42O4	453.3003	453.301	1.61	409.3111[M-H-CO2]^−, 391.2993[M-H-CO2-H2O]^−, 363.2662[M-H-CO2-H2O-CO]^−, 295.2077[M-H-C8H14O3]^−, 279.2118[M-H-C8H14O4]^−	Ursonic acid^b	ME, EE, HE
87	46.798	250	C39H54O6	617.3835	617.3848	2.04	279.2328[M-H-C21H22O4]^−	Pyracrenic acid^b	ME, EE
90	35.341	n.d	C29H46O5	473.3279	473.3272	−1.37		Segetalic acid^b	ME, EE, WE
91	53.076	220	C32H50O7	545.3487	545.3484	−0.59	449.3426[M-H-CO-H2O]^−,	Hovenidulcigenin B^b	ME, EE, HE
							423.3303[M-H-C3H6O5]^−
92	56.79	223	C32H44O9	571.2894	571.2912	3.24		Ganoderic acid H^b	ME
93	59.704	220	C36H62O8	621.435	621.4372	3.52		3-O-Protocatechuoylceanothic acid^b	ME, EE, HE
94	59.737	n.d	C30H44O3	451.3221	451.3218	−0.73		Zizyberenalic acid^b	ME, EE, HE
100	37.589	n.d	C30H44O4	467.3173	467.3167	−0.89		Ganoderic acid TR^b	ME, EE, HE
101	34.342	n.d	C30H44O6	499.306	499.3065	1.02	545.3109[M-H+ HCOOH]^−, 455.3188[M-H-CO2]^−, 437.3034[M-H-CO2-H2O]^−	3-Dehydroxy ceanothetric acid^b	ME, EE
102	54.392	222	C30H48O3	455.353	455.3531	−0.29		Oleanolic acid^b	ME, EE, HE
103	44.45	n.d	C32H48O6	527.3378	527.3378	0.02		Ganoderic acid V^b	ME, EE, HE
104	45.866	n.d	C30H48O4	471.347	471.348	2.08		Maslinic acid^b	ME, EE
106	57.639	n.d.	C39H54O6	617.3839	617.3848	1.4		Helicterilic acid^b	ME, EE
108	50.912	n.d	C30H46O5	485.3265	485.3272	1.54		Abrisapogenol I^b	ME, EE, WE
114	15.323	n.d.	C29H44O6	487.3058	487.3065	1.46		Polygalic Acid^b	EE
120	38.577	n.d.	C29H46O4	457.3324	457.3323	−0.15		Platanic acid^b	ME, EE

^aCompared with literatures.

^bCompared with MSⁿ data, databases, and/or characteristic UV spectra.

^cCompared with an authentic standard.

^dME: methanol extract; EE: ethyl acetate extract; HE: hexane extract; WE: water extract.

Figure 1. Total ion chromatogram (TIC) in negative ion mode (A1, A2) and DAD chromatogram at 280 nm (B1, B2); A1, B1: 0–30 min, A2, B2: 30–70 min.

Figure 2. MS/MS spectra and fragmentation patterns of some typical phetochemicals of the peduncles of Hovenia acerba Lindl.: gentiobiose (A); citric acid (B); protocatechuic acid (C); ferulic acid (D); rutin (E); ampelopsin (F); catechin (G); ceanothetric acid (H); jujubogenin (I); ceanothic acid (J); and oleanolic acid (K).

Figure 2. Continued.

Table 3. Calibration curve equatuin and results from UPLC-QqQ-MS/MS for the major chemical constituents in HAP^a.

No.	Compounds	Calibration curve equation	R^2	LOD (ng/mL)	LOQ (ng/mL)	ME^b (μg/g DW)^c	EE^b (μg/g DW)^c	HE^b (μg/g DW)^c	WE^b (μg/g DW)^c
Organic acids
8	Malic acid	y = 2.8702x-82.7599	0.9991	3.34	12.4	3525.13 ± 147.76	0.25 ± 0.01	9.11 ± 0.13	2953.08 ± 55.84
9	Citric acid	y = 2.0931x-83.3326	0.9998	1.01	7.02	842.34 ± 27.82	nd	1.86 ± 0.06	1117.51 ± 51.70
67	Abscisic acid	y = 16.3631x-5.9695	0.9994	0.84	5.28	4.07 ± 0.79	0.65 ± 0.09	0.02 ± 0.01	3.62 ± 0.27
121	Quinic acid	y = 0.3444x-22.9252	0.9990	8.13	31.67	249.20 ± 24.44	nd	nd	217.44 ± 6.06
Phenolic acids
15	Protocatechuic acid	y = 5.2912x+ 53.3888	0.9996	1.77	10.7	0.85 ± 0.08	0.12 ± 0.01	nd	1.16 ± 0.08
43	Chlorogenic acid	y = 3.1295x-0.4390	0.9994	1.64	6.52	1.76 ± 0.15	nd	nd	2.15 ± 0.62
44	p-hydroxybenzoic acid	y = 4.6335x+ 67.8022	0.9990	1.97	9.65	1.64 ± 0.23	0.41 ± 0.03	nd	1.96 ± 0.06
57	Ferulic acid	y = 1.1594x+ 4.8152	0.9996	5.31	20.34	4.53 ± 0.26	1.04 ± 0.02	nd	3.91 ± 0.43
63	Isoferulic acid	y = 1.0930x+ 6.3236	0.9997	2.07	13.8	5.54 ± 0.56	0.81 ± 0.02	nd	3.83 ± 0.72
Flavonoids
21	Epigallocatechin	y = 6.4614x+ 145.3070	0.9987	1.29	4.59	6.28 ± 0.12	nd	nd	3.50 ± 0.43
22	Epicatechin	y = 1.1375x+ 27.5539	0.9996	1.43	9.19	12.43 ± 0.53	0.09 ± 0.04	nd	4.30 ± 0.15
46	Ampelopsin	y = 6.1266x+ 4.5482	0.9990	0.87	5.57	8.40 ± 0.61	0.06 ± 0.00	nd	4.34 ± 0.40
56	Taxifolin	y = 10.4115x-13.7785	0.9997	1.29	7.03	0.51 ± 0.05	0.03 ± 0.00	nd	0.30 ± 0.02
59	Rutin	y = 2.1339x+ 62.5134	0.9994	0.82	3.73	11.45 ± 1.50	nd	nd	6.95 ± 1.46
61	Myricetin	y = 14.8619x+ 4.1188	0.9994	0.53	4.34	0.12 ± 0.06	nd	nd	nd
69	Chrysin	y = 3.7884x+ 472.3353	0.9990	1.21	4.87	nd	3.36 ± 0.03	nd	nd
70	Apigenin	y = 14.7241x+ 73.3896	0.9985	1.5	8.28	nd	0.02 ± 0.01	nd	nd
109	Gallocatechin	y = 5.8138x+ 87.4753	0.9989	0.77	4.29	43.88 ± 2.63	0.03 ± 0.01	nd	28.93 ± 1.13
115	Naringenin	y = 15.6237x-51.3137	0.9998	1.24	5.66	nd	0.19 ± 0.00	nd	nd
119	Genistein	y = 3.2793x+ 691.4639	0.9992	0.63	4.17	nd	0.96 ± 0.02	nd	nd
122	Catechin	y = 0.2234x+ 37.3966	0.9998	14.66	73.49	75.14 ± 5.83	1.21 ± 0.05	nd	36.44 ± 5.19
123	Naringenin-7-O-glucoside	y = 8.6962x-116.7114	0.9987	0.21	2.73	1.70 ± 0.20	0.08 ± 0.01	nd	0.87 ± 0.12
124	Quercetin	y = 7.4711x+ 951.8437	0.9997	0.54	3.86	1.02 ± 0.40	0.09 ± 0.01	nd	nd

^aResults are expressed as mean ± standard deviation of three replicates.

^bME: methanol extract; EE: ethyl acetate extract; HE: hexane extract; WE: water extract.

^cDW: dry weight of material.

nd: none detected.