In vitro and in vivo metabolism of Cistanche tubulosa extract in normal and chronic unpredictable stress-induced depressive rats

Highlights

• The metabolic profile of CTE in normal and depressed rats has been evaluated.

• A total of 20 and 26 metabolites were identified from in vitro and in vivo metabolism.

• CTE was metabolized to aglycones and degradation products by both normal and CUS rats.

• The metabolic capability has been altered by disordered gut microbiota in CUS rats.

Abstract

Cistanche tubulosa, one species of Cistanches Herba, was recently confirmed to have antidepressant efficacy in chronic unpredictable stress (CUS) rats by restoring homeostasis of intestinal microbiota. In this paper, we aim to explore the metabolic profile of C. tubulosa in normal and CUS induced depressive model rats in vitro and in vivo. Using UPLC-Q-TOF-MS, the in vitro gastrointestinal metabolism of Cistanche tubulosa extract (CTE) was evaluated in both normal and CUS rats. At the same time, in vivo metabolism of CTE in normal and depressed rats were also investigated in rat urine and feces. A total of 20 and 26 metabolites were characterized from in vitro and in vivo metabolism in normal and CUS rats, respectively. CTE was metabolized to aglycones and degradation products of phenylethanoid glycosides (PhGs) and iridoid glycosides whether by normal or depressed rat intestinal microbiota in vitro. Phase II metabolites of aglycones and degradation products of PhGs and iridoid glycosides were the main metabolites in rat urine and feces. Additionally, the metabolic capability to generate secondary glycosides and aglycones in depressive rat intestinal microbiota was much weaker than that in normal rat intestinal microbiota, which was attributed to the disordered glycoside hydrolases produced by intestinal microbiota in CUS depressed rats. The results of this study laid the foundation for understanding the metabolic process and therapeutic mechanism of CTE's antidepressant property.

Introduction

Cistanches Herba is officially recorded as the dried succulent stems of Cistanche deserticola (Y. C. Ma) and C. tubulosa (Schrenk), which is used to treat kidney deficiency, impotence, female infertility, morbid leucorrhea, profuse metrorrhagia, and senile constipation [1]. Modern pharmacological studies have shown that Cistanches Herba possesses various biological activities such as anti-neurodegeneration, immunoregulation, and anti-inflammation [2,3]. Our previous investigations have verified that C. tubulosa extract (CTE), which are consisted of 48.6% phenylethanoid glycosides (PhGs), 6.9% iridoid glycosides, and 20.0% total saccharides, could markedly alleviate depressive symptoms of chronic unpredictable stress (CUS)-induced depressive rats by restoring homeostasis of gut microbiota [4]. Recent studies indicate that changes in the intestinal microbiota composition were associated with the development and progression of depression [5,6]. The relative abundances of the microbial genera were markedly disturbed in CUS depressive model rats compared with normal controls [7]. In depressed patients, the diversity and richness of intestinal microbiota were also significantly altered [8]. Moreover, various compounds including phenylethanoid glycosides (PhGs) and iridoid glycosides were considered as the main constituents of Cistanches Herba [2,3], which were easily metabolized into their secondary glycosides and aglycones including hydroxytyrosol (HT), 3,4-dihydroxyphenethyl glycoside, deglycosylated geniposidic acid etc. by human intestinal microbiota. These metabolites are more easily absorbed through the intestine and exert biological activity consistent with the prototype component [[9], [10], [11]]. Thus, we believe that during the occurrence and development of depression, the disturbance of intestinal microflora structure will inevitably affect the metabolism of oral traditional Chinese medicines (TCMs) in the gastrointestinal tract, in addition to affecting the physiological state of the host. Most of the existing metabolic data of Cistanches Herba come from metabolic studies on healthy animals [[12], [13], [14], [15]]. Therefore, it would be of more clinical significance to investigate the metabolic profile of CTE in the pathological state in elucidating its bioactive components and understanding the mechanism of action for its anti-depressive efficacy.

In the current study, we aim to characterize the metabolic profiles of CTE in both healthy and CUS-induced depressive model rats by ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS). Gastric juice, intestinal fluid, and microbiota of normal and depressive pathological rats are used to simulate the metabolic process of CTE in the gastrointestinal tract in vitro, independently and sequentially. In vivo metabolites are also elucidated after oral administration of CTE in normal and CUS rats. This study provides new insights into the metabolism and active metabolites of CTE for depression.