Shuting Zou,1,* Bin Xie,2,* Zhentao An,1 Fang Li,3 Li Cui,1 Zhenhai Zhang,1 Weiquan Bu,1 Dandan He1 

1Affiliated Hospital of Integrated Traditional Chinese and Western Medicine, Nanjing University of Chinese Medicine, Nanjing, Jiangsu, 210028, People’s Republic of China; 2Department of Gastroenterology, Nanjing Lishui District Hospital of Traditional Chinese Medicine, Nanjing, Jiangsu, 211200, People’s Republic of China; 3Department of Gastroenterology, Changshu Hospital Affiliated to Nanjing University of Chinese Medicine, Changshu, Jiangsu, 215500, People’s Republic of China

*These authors contributed equally to this work

Correspondence: Weiquan Bu, Affiliated Hospital of Integrated Traditional Chinese and Western Medicine, Nanjing University of Chinese Medicine, No. 100, Cross Street, Hongshan Road, Qixia District, Nanjing, Jiangsu, 210028, People’s Republic of China, Email buweiquan2008@126.com Dandan He, Affiliated Hospital of Integrated Traditional Chinese and Western Medicine, Nanjing University of Chinese Medicine, No. 100, Cross Street, Hongshan Road, Qixia District, Nanjing, Jiangsu, 210028, People’s Republic of China, Email happydandan007@163.com

Objective: The aroma of the aromatic class of traditional Chinese medicines can promote gastrointestinal peristalsis. This study aimed to explore the mechanisms by which volatile oil from Citri Reticulatae Pericarpium Viride (VOCRPV) alleviates slow-transit constipation (STC).
Methods: The main active ingredients in VOCRPV were determined by High-Performance Liquid Chromatography (HPLC). Due to poor stability, an olive oil solution was prepared to enhance the volatile oil’s stability. A mouse model of STC was induced using loperamide hydrochloride. The mice’s body weight was monitored weekly. The number of fecal pellets, fecal water content, and small intestinal propulsion rate were detected. The colon tissues were analyzed using HE staining. The serum content of gastrointestinal hormones was measured using the corresponding ELISA kit. The protein expressions of stem cell factor (SCF) and c-Kit in colon tissues were detected by Western blot and immunohistochemistry methods. The 16S rRNA gene sequencing was used to detect the intestinal flora.
Results: The contents of p-isopropyl toluene, γ-Terpinene, and d-Limonene were determined by HPLC. VOCRPV and its olive oil solution significantly enhanced body weight, increased the number of fecal pellets, improved fecal water content, and boosted small intestinal propulsion rate in mice with loperamide-induced STC, while also repairing colon mucosa damage. They also increased gastrin (Gas) and motilin (MTL) levels in treated mice, upregulated the expression of SCF and c-Kit proteins, and restored intestinal flora balance in STC mice.
Conclusion: VOCRPV could effectively alleviate STC, and olive oil enhances its therapeutic effect. VOCRPV alleviates STC by elevating Gas and MTL levels, activating the SCF/c-Kit signaling pathway, and modulating intestinal flora.

Keywords: slow-transit constipation, volatile oil from Citri Reticulatae Pericarpium Viride, olive oil, stem cell factor/c-Kit signaling pathway, intestinal flora