Ginsenoside Re: Its chemistry, metabolism and pharmacokinetics

Ginsenosides, the bioactive components of ginseng, can be divided into two major groups, namely 20(S)-protopanaxatriol (e.g. Re, Rg1, Rg2, and Rb3) and 20(S)-protopanaxadiol (e.g. Rb1, Rb2, Rc, and Rd). Biological and environmental factors may affect the content of ginsenosides in different parts of ginseng plant. Evidence from pharmacokinetic and metabolic studies of Re demonstrated that (1) the absorption of Re is fast in gastrointestinal tract; (2) Re may be metabolized mainly to Rh1 and F1 by intestinal microflora before absorption into blood; and (3) Re is quickly cleared from the body.


Background
Ginseng is a key herb in Chinese medicine, and has a wide range of therapeutic and pharmacological uses [1][2][3]. Panax ginseng is a slow growing perennial herb of the Araliaceae family usually cultivated in China, Japan, Korea and Russia, as well as in the United States and Canada. Ginseng root has been used as an oriental folk medicine for several thousand years [2,4]. It is a highly valued medicinal plant in the Far East that and also popular in the West in the past 20 years [2,[4][5][6][7].
In order to explore the pharmacological actions, mechanisms and clinical applications of ginseng, some researchers focused on purified individual ginsenosides rather than whole ginseng extracts [1,23]. Individual ginsenosides may have different characteristics in chemistry, metabolism, and pharmacokinetics. Ginsenoside Re (Re) belongs to 20(S)-protopanaxatriol group (Figure 1), and is a major ginsenoside in ginseng [7,10,16,22,[24][25][26][27]. Literature shows that Re exhibits multiple pharmacological activities via different mechanisms [12,16,28]. For example, in cardiovascular system, Re possesses negative effects on cardiac contractility and autorhythmicity, anti-arrhythmic and anti-ischemic effects, angiogenic regeneration activities and cardiac electrophysiological functions [28]. Xie et al. and Li et al. [13,[29][30][31][32] found that the quantity of Re in ginseng leaf and berry is much higher than in ginseng main root and suggested that ginseng leaf-stem could be a valuable source for Re. There have been other new findings in recent years. This article provides an overview of the recent advances in chemistry, metabolism and pharmacokinetics of Re.
Ginsenosides are the glycosides that contain aglycone with dammarane (except Ro). Ginsenosides ( Figure 2) are generally divided into two groups, namely the protopanaxadiol (PPD) and protopanaxatriol (PPT) ginsenoside groups. The sugar moities in the PPD group including Rb 1 , Rb 2 , Rc, Rd, Rg 3 and Rh 3 , attach to the 3position of dammarane-type triterpine, whereas the sugar moities in the PPT group including Re, Rf, Rg 1 , Rg 2 and Rh 1 , attach to the 6-position of dammaranetype triterpine. A ginsenoside possesses a rigid four trans-ring steroid skeleton with a modified side chain at C-20 [1,17,[36][37][38][39]. The chemical structures of ginsenosides are different from each other in the number, linkage position and type of sugar [17,39]. During extraction, sugar moieties of ginsenosides may be cleaved by acid hydrolysis or endogenous glycosidases to give corresponding aglycones [25,36]. PPD and PPT may rearrange into panaxadiol and panaxatriol, respectively, to provide artificial ginsenoside products. Kang et al. [40] showed that PPD and PPT ginsenoside groups had different bioactivities, even opposite effects. Recently, Zhu et al. [41] found six new PPT-type ginsenosides extracted from the P. ginseng root, named Re 1 to Re 6 (compounds 1-6) respectively, along with ten other known PPT ginsenosides.

Content of Re in ginseng
The biological and environmental factors that may affect the quantity and quality of ginsenosides in ginseng [14,35] include the species, age, part of the plant, season of harvest, method of cultivation, and means of preservation. For example, the content of Re, Rg 1 and Rd is higher in the wild P. ginseng roots than in the cultivated ginseng roots, while the content of Rc, Rb 2 and Rb 1 is lower in the wild P. ginseng roots than in the cultivated ones. These differences in content of ginsenosides might affect their biological and pharmacological properties. Root ginsenoside content depends on the age of ginseng plant. For example, the plants younger than four years of age are considered unsuitable for harvest due to their low ginsenoside content [35,[42][43][44]. Lim et al. [35] determined the genotypes and environmental factors affecting the ginsenoside content among eight wild populations of P. quinquefolius. The influence of genotypes and environment on ginsenoside content varies among different types of ginsenosides. Specifically, the Re content varies with populations but not locations, whereas Rb 1 , Rc and Rb 2 only varies with locations, and Rg 1 and Rd varies with both. Ginsenoside levels are decreased, while ginseng growth is increased, at an intensively managed garden location. The content and composition of ginsenosides vary with other environmental conditions such as the type of soil, temperature, light intensity and water content [45].
Using high pressure microwave-assisted extraction (HPMAE) and high-performance liquid chromatography (HPLC) coupled with evaporative light scattering detection (ELSD), i.e. HPMAE HPLC-ELSD, Qu et al. [46] studied the effects of different parts and age of P. quinquefolius on the content of 12 ginsenosides, namely Rg 1 , Re, Rf, Rg 2 , Rh 1 , Rb 1 , Rc, Rb 2 , Rb 3 , Rd, Rh 2 and F 11 . The study ranked the parts of five-years-old P. quinquefolius in terms of total content of these 12 ginsenosides in a descending order: leaf, root-hair, rhizome, main root and stem, suggesting that the leaf could be a better source for ginsenosides, as compared with other parts of ginseng plant. It also found that in ginseng roots, the content of Re and Rb 1 , the major ginsenosides, increase with age of the plant.
In a comparative study on the quality of Tongrentang Red Ginseng and Korean Red Ginseng, Wu et al [47] found that the content of Re, Rg 1 and Rb 1 in the Tongrentang Red Ginseng is less than the content in the Korean Red Ginseng.
Another extensive study [48] performed a quantitative analysis of Re, Rb 1 and Rg 1 in P. quinquefolius berry and flower sampled in various months throughout the year, by enzyme-linked immunosorbent assay (ELISA). The P. quinquefolius flower had higher content of Re, Rb 1 and Rg 1 and the lowest content of Re in the berries harvested in September [48]. To analyze the Re content in P. quinquefolius berry pulp extracts, Morinaga et al. [49] performed a new Eastern blot technique with anti-Re monoclonal antibody, and confirmed that the content of Re varies from part to part in the plant.
Lee et al. [50] reported the variations in the ginsenoside profiles of ginseng landraces in Korea. They found that the P. ginseng wild population exhibits three types of ginsenoside profiles affected by genetic and environmental factors.

Metabolism and pharmacokinetics
Re has recently been studied extensively [12,13,30]; however, little is known about the metabolic and pharmacokinetic profiles.

Absorption
After oral administration, Re is in contact with the gastriointestinal fluids containing gastric acids and gastric enzymes, intestinal enzymes, and colonic bacteria [51,52]. Li et al. [23,53] studied the pharmacokinetic parameters and absolute bioavailability of Re, R 1 , Rg 1 , Rd, and Rb 1 after oral or intravenous administration of total notoginsenosides. Main pharmacokinetic parameters of these constituents were determined by Drug and Statistics (DAS) for Windows pharmacokinetics software. The results showed that Re, R 1 , Rg 1 , Rd and Rb 1 reached peak concentration in plasma within about 45 minutes after oral administration of total panax notoginsenoside (TPNG) powder in rats, suggesting a rapid absorption of ginsenosides in gastrointestinal tract. The absolute bioavailability of Re was 7.06% [53]. To confirm the rapid absorption finding, Joo et al. [27] conducted a pharmacokinetic study using ICR mice and ultra performance liquid chromatography-electrospray ionization-mass spectrometry (UPLC-ESI-MS) analytical method. This pharmacokinetic study [27] revealed that the time to reach the peak plasma concentration after oral administration was 0.4 ± 0.2 hour. The data also showed that the oral bioavailability was 0.19-0.28%. Qi et al. [54] found that the oral bioavailability of PPD ginsenosides (Ra 3 , Rb 1 , Rd, Rg 3 and Rh 2 ) and PPT ginsenosides (Rg 1 , Re, Rh 1 , and R 1 ) was less than 5% and PPT ginsenosides had better bioavailability, possibly due to the faster degradation of PPD ginsenosides.

Metabolism and biotransformation
Han et al. [55] showed that PPT ginsenosides are hydrolyzable to Rh 1 under mild acidic conditions. Tabaw et al. [56] found that two degradation products of the PPT ginsenosides, Rh 1 and F 1 could reach the systemic circulation in humans in addition to compound-K resulting from the stepwise deglycosylation of PPD ginsenosides. Bae et al. [57] further confirmed that the PPT (Re and Rg 1 ) could be metabolized mainly to Rh 1 and G-F1 in the gastrointestinal tract by intestinal microflora, before absorption into the blood. Chi and Ji et al. [58] tested the biotransformation of Re and Rb 1 by cell extracts from various food-grade edible microorganisms. As shown in Figure 3, Re was transformed into Rh 1 via Rg 2 by Bif. sp. Int57 and Bif. sp. SJ32; A. niger transformed Re into Rh 1 via Rg 1 ; A. usamii transformed Re into Rg 2 . However, Rb 1 was transformed into compound-K and Rh 1 by different pathways [58].
Metabolic research of Re in animals was also reported [59]. Six SD rats were used and divided into three groups. Feces were collected at 12, 24, 36, 48 and 60 hours after oral administration of Re (100mg/kg). Six metabolites of Re were detected in the feces of rat. The structures of the metabolites were identified as 20(S)ginsenoside Rg 2 , 20(S)-ginsenoside Rh 1 , 20(R)-ginsenoside Rh 1 , ginsenoside F 1 , 3-oxo-ginsenoside Rh 1 and PPT. The metabolic pathways of Re in animals were similar to those in humans [59].
A similar metabolic study was also carried out in vivo with HPLC coupled with electrospray ionization and quadrupole time-of-flight tandem mass spectrometry (HPLC-ESI-TOF-MS/MS) [60]. The rat urine samples were collected and pretreated through C(18) solid-phase extraction cartridges prior to analysis. As a result, eleven and nine metabolites together with Re were detected and identified in rat urine after oral and intravenous administration, respectively. Oxidation and deglycosylation were found to be the major metabolic processes of the constituent in rat, indicating that a large part of the intact ginsenosides was metabolized and transformed to ginsenosides with more biological effects in the gastrointestinal tract [52]. PPT ginsenosides, such as Re and Rg 1 , were mainly converted to Rh 1 and F 1 and then to corresponding aglycones [51,56].

Elimination
Xia et al. [38] applied a developed and validated liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS) method to detect Re, Rg 1 , Rd, Rb 1 and ophiopogonin D in rat plasma. Re and Rg 1 were eliminated quickly from the body. The pharmacokinetic behaviors of Rd and Rb 1 were significantly different from those of Re and Rg 1 in rat. Joo et al. [27] found that Re was rapidly cleared from the body within 0.2 ± 0.03 hour for male mice and 0.5 ± 0.08 hour for female mice after intravenous administration. They also found that ginseng berry extract exhibited a superior oral absorption of Re as compared to orally fed Re, suggesting that ginseng berry extract may be of choice for Re intake [27].
The plasma concentrations of Re and Rg 1 were determined and the pharmacokinetic parameters were calculated after intravenous Shenmai injection in ten volunteers [61]. The study found the distribution and elimination of Re and Rg 1 to be rapid after intravenous injection; and the pharmacokinetic characteristics could be fitted to the two-compartment model of pharmacokinetics.

Conclusion
Multiple biological and environmental factors affect the quantity and quality of ginsenosides in ginseng parts. Studies on Re demonstrate that (1) the absorption of Re is quick in rats; (2) PPT, Re and Rg 1 , are likely to be metabolized to Rh 1 and F 1 by intestinal microflora before absorption into the blood; and (3) Re can be quickly eliminated from the body.

Competing interests
The authors declare that they have no competing interests.