Pharmacological Effects and Molecular Mechanism of Synephrine Hydrochloride

Quan QUAN, Jingchao WANG, Mingxinzhi WANG, Hai CHEN, Anqi WANG, Chenghao JIN

College of Life Science and Biotechnology, Heilongjiang Bayi Agricultural University, Daqing 163319, China

Abstract Synephrine hydrochloride, an alkaloid compound extracted from immature bitter orange, has many pharmacological effects such as anticancer effect, antioxidant effect and antidepression effect. In this paper, the pharmacological effects and mechanism of synephrine hydrochloride were reviewed in order to provide a theoretical basis for the development and application of synephrine hydrochloride.

Key words Synephrine hydrochloride, Pharmacological effects, Anticancer, Antioxidation, Antidepression

1 Introduction

Immature bitter orange, also known as Fructus Aurantii Immaturus, is a dried young fruit of lime or sweet orange in the Rutaceae family. It was first published inShennongMateriaMedica[1]. It has a long medicinal history and has the effects of breaking qi, eliminating stagnation, and eliminating phlegm. It is mainly produced in Sichuan, Jiangxi, Fujian, Jiangsu and other places[2], and has been included in thePharmacopoeiaoftheRepublicofChinain 2020[3]. Synephrine hydrochloride is an alkaloid active component extracted from immature bitter orange. Its molecular formula is C9H13NO2. HCl, its molecular weight is 203.67, its melting point is 147-150 ℃, and it is white fine powder at normal temperature[4]. Studies have shown that synephrine hydrochloride has pharmacological effects such as anticancer effect, antioxidant effect and antidepression effect[5]. In this paper, the pharmacological effects and mechanism of synephrine hydrochloride in recent years were reviewed in order to provide a theoretical reference for further research and development of synephrine hydrochloride.

2 Anticancer effect and molecular mechanism

Cancer is a disease caused by many factors, which has the characteristics of infinite proliferation, metastasis and strong invasiveness. Epidemiological investigation shows that nearly 10 million people die of cancer every year, which seriously threatens human life and health[6]. At present, the commonly used chemotherapy drugs have some disadvantages, such as large toxic and side effects, long treatment cycle and high price. Therefore, looking for an efficient, safe and inexpensive natural anticancer drug is a hot spot in drug research today[7]. It is found that synephrine hydrochloride, as a natural medicine, has good anti-tumor effect, low toxicity and side effects and low cost.

Xuetal.[8]extracted synephrine hydrochloride from dried tangerine peel, and determined the effect of synephrine hydrochloride on the viability of esophageal cancer KYSE30 cells and KYSE270 cells by CCK-8 experiment. The results showed that synephrine hydrochloride could inhibit the viability of KYSE30 cells and KYSE270 cells in a dose-dependent manner. When the concentration of synephrine hydrochloride was 20 μmol/L, the inhibition rates of synephrine hydrochloride on the two kinds of cells were 71.1%±5.8% and 75.7%±6.2%, respectively. Transwell test was used to detect the effect of synephrine hydrochloride on migration and invasion of two kinds of esophageal cancer cells. The results showed that synephrine hydrochloride significantly inhibited the migration and invasion of KYSE30 cells and KYSE270 cells, with the inhibition rates of 73.3%±7.5% and 75.3%±3.4%, respectively, and had no obvious toxic and side effects on normal cells. The inhibitory effect of synephrine hydrochloride on proliferation and migration of esophageal squamous cell carcinoma was further detected by Western blot. The results showed that synephrine hydrochloride could significantly up-regulate the expression of E-cadherin and down-regulate the expression of Vimentin, p-ERK, p-AKT and Galectin-3. These results indicate that synephrine hydrochloride may inhibit the proliferation of esophageal squamous cell carcinoma by inhibiting AKT and ERK signaling pathways, and inhibit the migration and invasion of esophageal squamous cell carcinoma by regulating the expression of epithelial-mesenchymal proteins.

Wu Yaosongetal.[9]detected the inhibitory effect of synephrine hydrochloride on the viability of human gastric cancer SGC-7901 cells by MTT assay. The results showed that the viability of SGC-7901 cells was significantly inhibited after treated with synephrine hydrochloride for different time (24, 48 and 72 h). The effect of synephrine hydrochloride on apoptosis of SGC-7901 cells was detected by Annexin V-FITC/PI double staining assay and flow cytometry. The results showed that the apoptosis of SGC-7901 cells was significantly induced after 72 h treatment with synephrine hydrochloride, and the apoptosis rate was up to 40.12%. Furthermore, the effect of synephrine hydrochloride on SGC-7901 cell cycle was detected by flow cytometry. The results showed that the percentage of G1/G0cells increased significantly from 52.19% to 70.45% after treated with synephrine hydrochloride for 72 h. These results indicate that synephrine hydrochloride can induce apoptosis and cycle arrest of gastric cancer cells, and then inhibit the proliferation of gastric cancer cells.

Parketal.[10]detected the inhibitory effect of synephrine hydrochloride on human lung cancer A549 cells by MTT assay. The results showed that the viability of A549 cells was significantly inhibited after treated with synephrine hydrochloride for 24 h, and the median inhibitory concentration (IC50) was 220 μg/mL. Apoptosis and the expression of apoptosis-related proteins were detected by Annexin V-FITC/PI double staining assay and Western blot assay. The results showed that synephrine hydrochloride could induce apoptosis of A549 cells by up-regulating the expression of Bax caspase-3 and cleaved-PARP and down-regulating the expression of Bcl-2. The effects of synephrine hydrochloride on cell cycle of A549 cells were detected by flow cytometry and Western blot. The results showed that synephrine hydrochloride could significantly down-regulate the expression of Cyclin B1, cdc 25c and cdc 2, and up-regulate the expression of p21 protein, thus inducing A549 cell cycle arrest in G2/M phase. These results indicate that synephrine hydrochloride can inhibit the proliferation of lung cancer cells by inducing apoptosis and cycle arrest.

Shenetal.[11]extracted synephrine hydrochloride from immature bitter orange, and detected the inhibitory effects of synephrine hydrochloride and other compounds on the viability of human breast cancer MCF-7 cells and human lung cancer HCC827 cells by MTT assay. The results showed that the viability of MCF-7 cells and HCC827 cells was significantly inhibited by synephrine hydrochloride after treated with synephrine hydrochloride for 24 h. The expression of immune activity related factors was detected by quantitative real-time PCR (qRT-PCR) assay, and it was found that synephrine hydrochloride could up-regulate the mRNA expression of inducible nitric oxide synthase (iNOS), tumor necrosis factor α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6). The apoptosis-inducing effect of synephrine hydrochloride on MCF-7 cells and HCC827 cells was detected by Western blot. The results showed that synephrine hydrochloride could significantly up-regulate the expression of p-JNK, p-p38 and p-p65 proteins. These results indicate that synephrine hydrochloride can induce apoptosis of tumor cells by regulating MAPK signaling pathway, thus inhibiting the viability of breast cancer MCF-7 cells and lung cancer HCC827 cells.

3 Antioxidant effect

The antioxidant system of human body is a complex system, which can effectively inhibit the oxidation reaction of free radicals, and its mechanism is to directly act on free radicals or indirectly consume substances that are easy to produce free radicals. It is found that synephrine hydrochloride has good antioxidant effect[12].

Most fruits, such as litchi, are prone to peel browning after picking, and fat metabolism and oxidation can affect litchi peel browning. Heetal.[13]detected the effect of synephrine hydrochloride on various indexes in the process of litchi peel browning by spectrophotometry. The results showed that synephrine hydrochloride could significantly reduce the browning index and respiration rate of litchi fruit. The content of MDA and H2O2in litchi peel after picking was determined by thiobarbituric acid (TBA). The results showed that the content of MDA and H2O2in litchi peel of experimental group was significantly lower than that of control group. Further study showed that synephrine hydrochloride treatment increased the bilirubin content and the ratio of unsaturated fatty acids to saturated fatty acids in litchi peel, and significantly up-regulated the expression levels of delta-12 fatty acid desaturase FAD2 and omega-3 fatty acid desaturase FAD3 related to the formation of unsaturated fatty acids. These results indicated that synephrine hydrochloride could prolong the browning process of litchi peel by inhibiting lipid oxidation and regulating fat metabolism. Jiao Shirong[14]etal.detected the scavenging ability of synephrine hydrochloride on free radicals and the inhibitory effect on lipid peroxidation of mouse heart, liver and kidney homogenate by high performance liquid chromatography. The results showed that synephrine hydrochloride had strong scavenging ability on hydroxyl radical, superoxide anion radical and DPPH radical, and had good antioxidant effect. The effects of synephrine hydrochloride on renal antioxidant capacity and islet morphology in diabetic mice were detected by spectrophotometry. The results showed that the levels of blood glucose and malondialdehyde (MDA) decreased significantly, while the content of glutathione (GSH) increased significantly in diabetic model mice treated with synephrine hydrochloride. The effect of synephrine hydrochloride on liver antioxidant capacity of diabetic mice was detected by ferrous ion catalysis. The results showed that synephrine hydrochloride significantly inhibited glutathione peroxidase activity and NO content, and significantly increased the activity of oxidase dismutase. These results show that synephrine hydrochloride has good antioxidant effect.

4 Antidepression effect

Immature bitter orange can exert antidepression effect in many ways, including increasing the expression level of glucocorticoid receptor (GR) mRNA in hippocampus and inhibiting the hyperactivity of hypothalamus-pituitary-adrenal cortex axis; improving the expression level of brain-derived neurotrophic factor (BDNF) mRNA in hippocampus and cortex; improving the expression and release of neurotransmitters, such as 5-hydroxytryptamine, norepinephrine and dopamine; regulating gastrointestinal hormones and increasing gastrointestinal motility, such as increasing the secretion of gastrin and motilin[15].

Wang Jingjing[16]detected the antidepression effect of synephrine hydrochloride on chronic stress depression model rats by high performance liquid chromatography-fluorescence detection (HPLC-FLD). It was found that synephrine hydrochloride could significantly increase the content of 5-hydroxytryptamine (5-HT) and corticosterone in rats. Enzyme-linked immunosorbent assay (ELISA) was used to detect the effect of synephrine hydrochloride on serum corticosteroid in rats. It was found that the corticosteroid content in model group was significantly higher than that in control group. The ultrastructural changes of hippocampus induced by synephrine hydrochloride were detected by electron microscope. Compared with the control group, the cell membrane of hippocampus in model group showed serious folds, nucleolar pyknosis, and decrease of mitochondria and other organelles. Immunohistochemical technique and qRT-PCR were used to detect the expression of BDNF and GR in rat hippocampus. It was found that the number of BDNF and GR positive cells and the expression level of mRNA in synephrine hydrochloride treatment group decreased significantly. Xu Yingetal.[17]detected the intervention effect of synephrine hydrochloride on depression model rats by reverse transcription-polymerase chain reaction (RT-PCR), and found that synephrine hydrochloride had a good relieving effect on depressive symptoms in experimental rats, and its mechanism was related to improving gastric motility and up-regulating the expression level of related mRNA in cerebral cortex and hippocampus. Xie Qietal.[18]detected the effect of synephrine hydrochloride on the changes of endogenous metabolites in serum of rats by GC-MS, and found that synephrine hydrochloride could improve depressive symptoms of rats by regulating amino acid metabolism. These results indicate that synephrine hydrochloride plays an antidepression role by regulating the expression levels of BDNF and GR.

5 Outlook

Synephrine hydrochloride, as an alkaloid, has a wide range of pharmacological effects, but its specific molecular mechanism has not been deeply explored. Therefore, in the follow-up study, the pharmacological mechanism of synephrine hydrochloride should be studied more comprehensively and deeply at the cellular, molecular and animal levels, in order to provide a theoretical basis for further development and utilization of synephrine hydrochloride.