Biotransformation of Rare Protopanaxadiol Saponin by Human Intestinal Microflora†

doi:10.7503/cjcu20180812

Abstract

Abstract:

The rapid resolution liquid chromatography-quadrupole time-of-flight mass spectrometry(RRLC-Q-TOF MS) method and ultra high performance liquid chromatography-triple quadruple mass spectrometry(UPLC-QQQ MS) method were applied for qualitative and quantitative analyzing the biotransformation process of rare protopanaxadiol saponin Rd, Rg₃, F₂, CK and Rh₂ under intestinal micro flora in vitro. The rare protopanaxadiol saponin Rd, F₂, Rg₃, CK, Rh₂ and intestinal micro flora were cultured in vitro at 37 ℃ under anaerobic condition. The mass spectrometrics detection with electrospray ionization under negative ion mode was applied for identifying the metabolites, monitoring content variation, and matching metabolic pathways. The results show ginsenoside Rd is mainly metabolized into five products of F₂, Rg₃, CK, Rh₂ and PPD. Ginsenoside F₂ is mainly metabolized into two products of CK and PPD. Ginsenoside Rg₃ is mainly metabolized into two products CK and PPD. Ginsenosides CK and Rh₂ are both mainly metabolized into one product PPD. Ginsenosides Rd, F₂ and Rg₃ are metabolized completely into metabolic products, but ginsenosides CK and Rh₂ are not, under human intestinal micro flora condition in vitro. The results also indicated that deglycosylation reaction was the major metabolic pathway of rare protopanaxadiol saponin under the human intestinal microflora condition. And that monoglycosides and aglycone may be the material basis for rare protopanaxadiol saponin to exert its efficacy in human body.

Key words: Rare protopanaxadiol saponin, Human intestinal micro flora, Biotransformation, Liquid chromatography-mass spectrometry

CLC Number:

O657

TrendMD:

HAN Mingxin, LI Fangtong, ZHANG Yan, DAI Yulin, ZHENG Fei, YUE Hao. Biotransformation of Rare Protopanaxadiol Saponin by Human Intestinal Microflora^†[J]. Chem. J. Chinese Universities, 2019, 40(7): 1390.

Figures/Tables 5

References 20

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Sample	Compd.	t_R/min	M_w	MS, m/z	MS/MS fragment ion, m/z	Relative error/ (ng·mL^-1)	Product
M1	Rd	26.187	991.5478	991.5443[M+HOOCH-H]^-	783.4929[M-H-Glc]^-,	3.53	F₂, Rg₃, CK,
					621.4354[M-H-2Glc]^-,		Rh₂, PPD
					459.3819[M-H-3Glc]^-
M2	F₂	31.421	829.4949	829.4947[M+HOOCH-H]^-	621.4308[M-H-Glc]^-,	0.24	CK, PPD
					459.3821[M-H-2Glc]^-
M3	Rg₃	32.693	829.4949	829.4884[M+HOOCH-H]^-	621.4311[M-H-Glc]^-,	7.84	Rh₂, PPD
					459.3824[M-H-2Glc]^-
M4	CK	35.411	667.4421	621.4369[M-H]^-/	459.3829[M-H-Glc]^-	2.70	PPD
				667.4439[M+HOOCH-H]^-
M5	Rh₂	36.002	667.4421	621.4373[M-H]^-/	459.3814[M-H-Glc]^-	1.65	PPD
				667.4432[M+HOOCH-H]^-
M6	PPD	40.201	505.3893	459.3878[M-H]^-/	221.0679[M-CHO-Glc]^-	0.79	—
				505.3897[M+HOOCH-H]^-

Sample	Compd.	t_R/min	M_w	MS, m/z	MS/MS fragment ion, m/z	Relative error/ (ng·mL^-1)	Product
M1	Rd	26.187	991.5478	991.5443[M+HOOCH-H]^-	783.4929[M-H-Glc]^-,	3.53	F₂, Rg₃, CK,
					621.4354[M-H-2Glc]^-,		Rh₂, PPD
					459.3819[M-H-3Glc]^-
M2	F₂	31.421	829.4949	829.4947[M+HOOCH-H]^-	621.4308[M-H-Glc]^-,	0.24	CK, PPD
					459.3821[M-H-2Glc]^-
M3	Rg₃	32.693	829.4949	829.4884[M+HOOCH-H]^-	621.4311[M-H-Glc]^-,	7.84	Rh₂, PPD
					459.3824[M-H-2Glc]^-
M4	CK	35.411	667.4421	621.4369[M-H]^-/	459.3829[M-H-Glc]^-	2.70	PPD
				667.4439[M+HOOCH-H]^-
M5	Rh₂	36.002	667.4421	621.4373[M-H]^-/	459.3814[M-H-Glc]^-	1.65	PPD
				667.4432[M+HOOCH-H]^-
M6	PPD	40.201	505.3893	459.3878[M-H]^-/	221.0679[M-CHO-Glc]^-	0.79	—
				505.3897[M+HOOCH-H]^-

Ginsenoside	Quantitation transition	Linear regression equation	Correlation coefficient(r²)	Linear range/ (μmol·mL^-1)	LOD/ (μmol·mL^-1)	LOQ/ (μmol·mL^-1)
Rd	991.5457, 783.4927	y=156278.26x+17382.41	0.995	0.58—142.82	0.032	0.093
F₂	829.4947, 621.4297	y=195432.18x+32126.16	0.991	0.62—81.51	0.009	0.034
Rg₃	829.4932, 621.4307	y=176368.23x+37525.19	0.992	0.32—72.89	0.007	0.032
CK	667.4435, 459.3828	y=374215.12x+24656.32	0.998	0.72—186.14	0.033	0.094
Rh₂	667.4371, 459.3842	y=283754.67x+26396.29	0.997	0.52—168.37	0.031	0.091
PPD	505.3887, 221.0676	y=127941.39x+41259.15	0.994	0.19—192.33	0.035	0.042

Ginsenoside	Quantitation transition	Linear regression equation	Correlation coefficient(r²)	Linear range/ (μmol·mL^-1)	LOD/ (μmol·mL^-1)	LOQ/ (μmol·mL^-1)
Rd	991.5457, 783.4927	y=156278.26x+17382.41	0.995	0.58—142.82	0.032	0.093
F₂	829.4947, 621.4297	y=195432.18x+32126.16	0.991	0.62—81.51	0.009	0.034
Rg₃	829.4932, 621.4307	y=176368.23x+37525.19	0.992	0.32—72.89	0.007	0.032
CK	667.4435, 459.3828	y=374215.12x+24656.32	0.998	0.72—186.14	0.033	0.094
Rh₂	667.4371, 459.3842	y=283754.67x+26396.29	0.997	0.52—168.37	0.031	0.091
PPD	505.3887, 221.0676	y=127941.39x+41259.15	0.994	0.19—192.33	0.035	0.042