The effect of gut microbiota on the activity of the HPA and HPG axes and ECS is a short-term or long-lasting process remains unclear in rodents. However, the extant studies focused only on its short-term effect on the HPA activity because there is lack of reliable biomarkers characterizing short-term activity of the HPG axis and ECS and long-term activities of the three endocrine systems. The endogenous levels of aldosterone (ALD), 11-dehydrocorticosterone (11-DHC), estradiol (E2), estrone (E1), androstenedione (A4), dihydrotestosterone (DHT), corticosterone (CORT), dehydroepiandrosterone (DHEA), testosterone (T), progesterone (P), N-arachidonoyl ethanoamide (AEA) and 1-arachydonoyl glycerol (1-AG) in hair and plasma are the potential long-term and short-term biomarkers of the three systems. This study aimed to develop the sensitive and selective methods for simultaneous quantitation of the twelve compounds in rodent's hair and plasma. Then the methods were used to explore the differences in the hair levels of the twelve compounds between the mice in XZ group possibly having gut microbiota with more diversity and SPF group possibly having gut microbiota with less diversity and the inter-group differences in the plasma levels in the response to 1-h restraint stress. The methods were based on high performance liquid chromatography tandem mass spectrometry with atmospheric pressure chemical ionization in positive mode. The methods were adopted for 20 mg hair and 100 μL plasma, respectively. Hair samples were incubated in methanol at 40 ℃ for 24 h, and were performed by solid phase extraction. Plasma samples were implemented by liquid-liquid extraction with ethyl acetate. The methods showed limit of quantification at 0.06-1.3 pg/mg and 0.03-0.6 ng/mL and recovery ranging between 87.7-115.1 % and 86.1-114.6 % for all compounds in rodent's hair and plasma, and the intra-day and inter-day coefficients of variation less than 15 %, and good freeze/thaw and short-term stability. The present methods also had good reliability as demonstrated by the sex difference in the testosterone levels in hair and plasma. The inter-group comparison revealed that the mice in XZ group showed significantly higher hair levels than those in SPF group for 1-AG and most of hormones except for T and P. The non-stressed female mice in SPF showed significantly higher plasma levels than those in XZ for AEA and most of hormones except for E2, A4, DHT, T and 1-AG, but there were no inter-group differences for the stressed mice except for DHEA and 11-DHC and for the non-stressed male mice. Additionally, the stressed mice showed significantly higher corticosterone level in plasma than controls for male and female mice in XZ and male mice in SPF, but it was not true for female mice in SPF.

中文翻译：

---

LC-APCI + -MS / MS方法分析了具有不同肠道菌群的小鼠血浆和毛发中的十种激素和两种内源性大麻素。

在啮齿类动物中，肠道菌群对HPA和HPG轴以及ECS的活动是短期还是长期的影响尚不清楚。但是，现有研究仅集中于其对HPA活性的短期影响，因为缺乏可靠的生物标记物来表征HPG轴和ECS的短期活性以及三个内分泌系统的长期活性。内源性醛固酮（ALD），11-脱氢皮质酮（11-DHC），雌二醇（E2），雌酮（E1），雄烯二酮（A4），二氢睾丸酮（DHT），皮质酮（CORT），脱氢表雄酮（DHEA），睾丸激素（ T），头发和血浆中的孕酮（P），N-花生四烯酰基乙酰胺（AEA）和1-花生四烯酸甘油酯（1-AG）是这三种系统的潜在长期和短期生物标志物。这项研究旨在开发灵敏和选择性的方法，用于同时定量啮齿动物头发和血浆中的十二种化合物。然后使用这些方法探索XZ组小鼠肠道菌群多样性较高的小鼠和SPF组肠道菌群多样性较低的小鼠之间的十二种化合物的毛水平差异以及血浆水平的组间差异在对1小时约束压力的反应中。该方法基于高效液相色谱串联质谱法，并采用正压大气压化学电离。该方法分别用于20 mg头发和100μL血浆。将头发样品在40℃的甲醇中孵育24小时，然后通过固相萃取进行。通过用乙酸乙酯进行液-液萃取来实施血浆样品。这些方法显示出啮齿动物头发和血浆中所有化合物以及当日和日间的定量限分别为0.06-1.3 pg / mg和0.03-0.6 ng / mL，回收率在87.7-115.1％和86.1-114.6％之间。日变化系数小于15％，并具有良好的冻融特性和短期稳定性。如通过头发和血浆中睾丸激素水平的性别差异所证明的，本方法还具有良好的可靠性。组间比较显示，对于1-AG和除T和P以外的大多数激素，XZ组的小鼠的毛发水平明显高于SPF组的毛发。 XZ中的AEA和大多数激素（除E2，A4，DHT，T和1-AG外）但是除了DHEA和11-DHC以及非应激雄性小鼠外，应激小鼠的组间差异均没有。此外，在XZ中雄性和雌性小鼠以及SPF中雄性小鼠的应激小鼠血浆中皮质酮水平明显高于对照组，但在SPF中雌性小鼠并非如此。