Identification of Polygonum orientale constituents using high-performance liquid chromatography high-resolution tandem mass spectrometry

Figures & data

Figure 1. MS and UV chromatograms of P. orientale extracts. (A) negative ionization mode; (B) positive ionization mode; (C) 330 nm.

Table 1. Peak identification of P. orientale extracts on chromatographic profiles in Figure 1.

Peak No.	Compounds identification	tR (mins)	Observed m/z	Calculated m/z	Molecular formula [M–H]^−	MS/MS fragments (m/z)	UV (λmax, nm)	Compounds
a	Isoorientin	10.076	447.0926	447.0933	C21H19O11	429 [M–H2O-H]^−	268, 350	11
b	Orientin	10.486	447.0924	447.0933	C21H19O11	327 [M–C4H8O4–H]^−	268, 350	10
c	Vitexin	11.407	431.0976	431.0978	C21H19O10	311 [M–C4H8O4–H]^−	268, 350
d	Isovitexin	11.612	431.0978	431.0978	C21H19O10	311 [M–C4H8O4–H]^−	269, 340
e	Cynaroside	12.021	447.0929	447.0933	C21H19O11	285 [M–C6H10O5–H]^−	266, 350	12
f	Catechin gallate	12.175	441.0811	441.0827	C22H18O10	289 [M–C7H4O4–H]^−	276
g	Rhamnetin-3-O-glucoside	12.482	477.1022	477.1038	C22H21O12	315 [M–C6H10O5–H]^−	270, 350
h	Flavonoid glycoside	12.943	447.0925	447.0933	C21H19O11	284 [M–C6H11O5–H]^−	266, 350
i	Quercitrin	13.097	447.0916	447.0933	C21H19O11	300 [M–C6H11O4–H]^−	258, 350	9
j	Afzelin	14.120	431.0974	431.0978	C21H19O10	284 [M–C6H11O4–H]^−	263, 350	8
k	Luteolin	16.270	285.0384	285.0405	C15H9O6	175 [M–C3O2–CH2CO–H]^−	276, 333
l	Unknown compound	16.782	329.0292	329.0303	C9H13O13	314 [M–CH3–H]^−	246, 367
m	Vanicoside C	17.857	821.2300	821.2298	C41H41O18	675 [M–C9H6O2–H]^−	315
n	Vanicoside B	18.881	955.2673	955.2666	C49H47O20	809 [M–C9H6O2–H]^−	315
o	Flavonoid	18.936	455.0976	455.0973	C23H19O10^a	440 [M–CH3+H]^−	280, 322
p	5,3′,4′-Trihydroxy-3-methoxy-6,7-methylenedioxyflavone	19.547	343.0442	343.0459	C17H11O8	328 [M–CH3–H]^−	328
q	Vanicoside A	19.956	997.2783	997.2772	C51H49O21	851 [M–C9H6O2–H]^−	315
r	Methoxyeugenol	20.314	193.0845	193.0870	C11H13O3	178 [M–CH3–H]^−	315
s	3,3′,4′,5,5′,8-Hexamethoxy-6,7-methylenedioxyflavone	21.496	447.1315	447.1286	C22H23O10^a	417 [M–CH2O+H]^+	280, 330	7
t	Gossypetin tetramethyl ether	21.901	373.0908	373.0929	C19H17O8	343 [M–CH2O–H]^−	280, 340
u	3,3′,5-Trimethoxy-4′,5,6,7′-bis(methylenedioxy)flavone	21.906	423.0711	423.0687	C20H17O9^a	408 [M–CH3+Na]^+	280, 340
v	5,5′-Dihydroxy-3,3′,4′,8-tetramethoxy-6,7-methylenedioxyflavone	22.008	419.1004	419.0973	C20H19O10^a	403 [M–O+H]^+	280, 350	5
w	3,3′,5,8-Tetramethoxy-4′,5′,6,7-bis(methylenedioxy)flavone	22.264	431.1007	431.0973	C21H19O10^a	401 [M–CH2O+H]^+	280, 340	6
x-1	Digicitrin	22.315	435.1323	435.1286	C21H23O10^a	405 [M–CH2O+H]^+	275, 330	1
x-2	Exoticin	22.418	463.1627	463.1599	C23H27O10^a	433 [M–CH2O+H]^+	275, 330	3
y	3,3′,5,6,7,8,-Hexamethoxy-4′,5′-methylenedioxyflavone	23.135	447.1315	447.1286	C22H23O10^a	417 [M–CH2O+H]^+	256, 345	2
z	5-Hydroxy-3,3′,6,7,8-pentamethoxy-4′,5′-methylenedioxyflavone	25.695	433.1161	433.1129	C21H21O10^a	403 [M–CH2O+H]^+	280, 350	4

^a [M + H]^+.

Figure 2. Chemical structures of isolated compounds 1–12 from P. orientale extracts.

Table 2. ¹H- and ¹³C NMR spectroscopic data for compounds 1–7 (δ in ppm).

Position	1^a		2^a		3^a		4^a		5^b		6^c		7^c
	^1H- NMR	^13C NMR	^1H- NMR	^13C NMR	^1H- NMR	^13C NMR	^1H- NMR	^13C NMR	^1H- NMR	^13C NMR	^1H- NMR	^13C NMR	^1H- NMR	^13C NMR
2		155.4		152.7		152.7		155.4		155.1		151.4		151.6
3		139.3		143.9		143.9		138.9		138.6		140.2		139.5
4		179.4		173.8		173.9		179.3		178.7		172.3		172.4
5		149.1		148.2		148.2		149.2		136.0		134.9		135.0
6		136.2		140.9		141.2		135.4		129.8		136.8		136.8
7		153.0		151.4		151.4		152.3		145.3		145.5		145.7
8		132.9		137.8		137.8		132.8		124.4		127.8		127.8
9		144.9		146.6		146.7		144.8		144.3		143.0		143.2
10		107.5		115.0		115.0		109.1		107.1		113.2		113.2
11									6.20 (2H, s)	103.4	6.20 (2H, s)	103.1	6.20 (2H, s)	103.2
1′		126.0		124.9		126.0		124.5		124.9		124.2		125.5
2′	7.40 (1H, d, 2.1)	104.9	7.52 (1H, d, 1.5)	108.7	7.51 (1H, s)	105.7	7.53(1H, d, 1.5)	107.4	7.22 (1H, d, 2.0)	103.6	7.37 (1H, d, 1.5)	108.3	7.37 (1H, s)	105.3
3′		152.1		143.5		153.1		143.5		153.0		143.0		152.8
4′		138.0		137.4		140.2		137.4		139.0		137.0		140.5
5′		149.3		149.1		153.1		149.2		150.7		148.7		152.8
6′	7.46 (1H, d, 2.1)	108.8	7.40 (1H, d, 1.5)	102.7	7.51 (1H, s)	105.7	7.41 (1H, d, 1.5)	102.9	7.29 (1H, d, 2.0)	109.7	7.23 (1H, d, 1.5)	102.1	7.37 (1H, s)	105.3
7′	6.07 (2H, s)		6.07 (2H, s)	102.1			6.10 (2H, s)	102.2			6.13 (2H, s)	101.7
OMe	3.89 (3H, s)	56.0	3.88 (3H, s)	56.6	3.90 (3H, s)	56.2 (2C)	3.88 (3H, s)	56.6	3.77 (3H, s)	55.8	3.78 (3H, s)	56.3	3.76 (3H, s)	55.9
	3.94 (9H, s)	60.3	3.94 (3H, s)	59.9	3.94 (12H, s)	60.0	3.96 (6H, s)	60.2	3.83 (3H, s)	59.9	3.85 (3H, s)	59.3	3.81 (3H, s)	55.9
	4.01 (3H, s)	61.1	3.97 (3H, s)	61.7	3.97 (3H, s)	61.0	3.98 (3H, s)	61.2	3.84 (3H, s)	60.0	3.91 (3H, s)	61.0	3.86 (9H, s)	59.5
	4.10 (3H, s)	61.2	3.98 (3H, s)	61.9	4.00 (3H, s)	61.7	4.11 (3H, s)	61.7	3.96 (3H, s)	61.3	4.00 (3H, s)	61.1	4.00 (3H, s)	60.2
		61.7	3.99 (3H, s)	62.0	4.09 (3H, s)	61.8		62.1						61.1
		62.1	4.09 (3H, s)	62.3		61.9								61.2
						62.3

^a Recorded in chloroform-d, 300, 75 MHz;

^b Recorded in DMSO-d_6, 600, 125 MHz;

^c Recorded in DMSO-d_6, 600, 75 MHz;.

Table 3. Effect of isolated compounds on NO production in BV2 cells, DPPH assay, and glutamate-induced toxicity in HT22 cells.

Compounds	IC50, μM^a
	NO	DPPH	Glutamate
1	9.7 ± 0.7	>100	>100
2	34.9 ± 4.4	>100	>100
3	14.3 ± 0.5	>100	>100
4	>100	>100	>100
5	35.8 ± 0.5	>100	>100
6	>100	>100	>100
7	11.4 ± 2.1	>100	>100
8	41.4 ± 1.4	>100	2.4 ± 2.1
9	>100	11.5 ± 0.4	40.4 ± 9.9
10	>100	9.7 ± 0.9	>100
11	>100	8.6 ± 0.1	>100
12	>100	14.5 ± 0.6	1.7 ± 0.6
Positive control^b	4.6 ± 0.1	26.8 ± 0.2	68.0 ± 2.0

^a IC50 value of each compound was defined as the concentration (μM) that caused 50% inhibition.

^b NO dexamethasone; DPPH curcumin; glutamate Trolox.