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Spectrum-Effect Relationship Between Serum HPLC Fingerprints and Activation of Blood Circulation and Removal of Blood Stasis by Chuanxiong

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Spectrum-Effect Relationship Between Serum HPLC Fingerprints and Activation of Blood Circulation and Removal of Blood Stasis by Chuanxiong Chen et al Tropical Journal of Pharmaceutical Research April 2021; 20 (4): 757-765 ISSN: 1596-5996 (print); 1596-9827 (electronic) © Pharmacotherapy Group, Faculty of Pharmacy, University of Benin, Benin City, 300001 Nigeria. Available online at http://www.tjpr.org http://dx.doi.org/10.4314/tjpr.v20i4.14 Original Research Article Spectrum-effect relationship between serum HPLC fingerprints and activation of blood circulation and removal of blood stasis by Chuanxiong Ling Chen, Wei Deng, Shirong Liu, Fengchao Li, Huimin Li, Guihua Jiang* College of Pharmacy, Chengdu University of Traditional Chinese Medicine, No. 1166 Liutai Avenue, Wenjiang District, Chengdu, 611137, China *For correspondence: Email: [email protected]; Tel: +86 18980923782 Sent for review: 26 November 2020 Revised accepted: 20 March 2021 Abstract Purpose: To study the active ingredients of Chuanxiong (CX) in promoting blood circulation and removing blood stasis. Methods: Blood-activating effects and serum HPLC fingerprints of CX extracts from different parts of China were studied and the spectrum-effect relationship between them was established by grey correlation analysis. Results: After treatment with CX extracted using different solvents, hemorheology indices were lower than those in the model group, and the effect of dichloromethane was better than that of other solvents (p < 0.05 or p < 0.01). There were 6 common peaks by fingerprint analysis. Peaks 1 - 5 were identified as senkyunolide I, senkyunolide H, senkyunolide A, coniferyl ferulate and ligustilide, respectively. Conclusion: Analysis of the spectrum–effect relationship indicates the contribution of the five components to the blood-activating effect of CX. The findings lend some scientific justification for using CX to remove blood stasis, and selection of quality evaluation indices for CX. Keywords: Chuanxiong, Blood-activating, Dichloromethane, Serum fingerprint, Senkyunolide I, Senkyunolide H, Senkyunolide A, Coniferyl ferulate, Ligustilide This is an Open Access article that uses a fund-ing model which does not charge readers or their institutions for access and distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0) and the Budapest Open Access Initiative (http://www.budapestopenaccessinitiative.org/read), which permit unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. Tropical Journal of Pharmaceutical Research is indexed by Science Citation Index (SciSearch), Scopus, International Pharmaceutical Abstract, Chemical Abstracts, Embase, Index Copernicus, EBSCO, African Index Medicus, JournalSeek, Journal Citation Reports/Science Edition, Directory of Open Access Journals (DOAJ), African Journal Online, Bioline International, Open-J-Gate and Pharmacy Abstracts INTRODUCTION main chemical components of CX are volatile oils, alkaloids, organic acids and phenols. Chuanxiong (CX) is the dried rhizome of Ligusticum chuanxiong Hort. CX is a well-known According to TCM theory, if the blood is clear, plant used in traditional Chinese medicine (TCM) then pain will be absent. Studies on the formulations which activates the blood circulation pharmacodynamic basis of the effects of CX and relieves pain [1]. Usually, it is employed with have focused primarily on chuanxiongzine, ferulic Angelica sinensis radix and Salvia miltiorrhiza for acid, ligustilide and senkyunolide with little or no the treatment of irregular menstruation, migraine, chuanxiongzine in the original CX or CX extracts. coronary heart disease, and angina [2-4]. The Studies [5-7] have shown that the main components in the plasma of rats after CX -------------------------------------------------------------------------------------------------------------------------------Trop J Pharm Res, April ----------------------2021; 20(4): 757 © 2021 The authors. This work is licensed under the Creative Commons Attribution 4.0 International License Chen et al administration are ferulic acid, ligustilide and National Institutes of Health, Bethesda, MD, senkyunolide, but chuanxiongzine is absent. Li USA) [11]. and colleagues [8] showed that endoesters were the main active ingredients of CX, and that CX Materials had a good effect on ischemic cardiac disease and cerebrovascular disease. Hence, the identity Twenty-two batches of CX were collected from of the main components of CX that have major producing areas in Sichuan Province, one pharmacodynamic effects is not known. batch of CX was collected from Fujian Province, and one batch of CX was collected from Shanxi We investigated the active ingredients of CX that Province, in China, and identified by Professor promote blood circulation and remove blood Yu-Ying Ma and Professor Gui-Hua Jiang stasis. The aim of the study was to determine the (College of Pharmacy, Chengdu University of “spectrum–effect relationship” between serum TCM, Chengdu, China). Information on these 24 high-performance liquid chromatography (HPLC), batches of CX is shown in Table 1. as well as the effect of CX on blood circulation and removal of blood stasis by grey correlation HPLC was carried out on a LC-2012 system analyses. (Shimadzu, Kyoto, Japan). An automatic hemorrheometer (SA-6000; Succeeder, Beijing, EXPERIMENTAL China) was used to assay blood samples. Methanol (HPLC - grade) was obtained from Ethical approval of the study protocol Thermo Fisher Scientific (Waltham, MA, USA). Ethanol, petroleum ether, dichloromethane, ethyl Animal experiments were undertaken with the acetate, N-butanol, and picric acid were approval of the Ethics Committee of Chengdu purchased from Chengdu Kelong Chemical University of Traditional Chinese Medicine Reagents (Chengdu, China). Z-ligustilide, (Chengdu, China). Animal treatments were levistilide A, senkyunolide I, senkyunolide A, conducted in strict compliance with the Guide for coniferyl ferulate, and senkyunolide H were the Care and Use of Laboratory Animals (US obtained from Chroma Biotechnology (Chengdu, China), and their purity was >95%. Table 1：Information on chuanxiong batches Batch Place of Altitude Latitude (E) Longitude (N) Time of number production collection CX1 Meishan, Sichuan 427.0 103º38′10.7″ 30º03′25.1″ 2017.04 CX2 Meishan, Sichuan 412.0 103º45′05.2″ 30º05′11.4″ 2017.04 CX3 Meishan, Sichuan 383.0 103º52′15.9″ 29º54′44.9″ 2017.04 CX4 Meishan, Sichuan 395.0 103º50′30.9″ 29º57′05.1″ 2017.04 CX5 Meishan, Sichuan 438.0 103º45′19.5″ 30º11′46.8″ 2017.04 CX6 Meishan, Sichuan 426.0 103º43′49.4″ 30º08′20.8″ 2017.04 CX7 Meishan, Sichuan 444.0 103º47′18.2″ 30º28′43.8″ 2017.04 CX8 Meishan, Sichuan 442.0 103º47′57.5″ 30º14′55.2″ 2017.04 CX9 Meishan, Sichuan 427.0 103º49′22.9″ 30º12′36.3″ 2017.04 CX10 Shifang, Sichuan 533.0 104°05′55.5″ 31°05′36.1″ 2017.04 CX11 Shifang, Sichuan 530.0 104°05′54.0″ 31°08′27.0″ 2017.04 CX12 Pengzhou, Sichuan 674.0 103°54′48.3″ 31°05′31.2″ 2017.05 CX13 Pengzhou, Sichuan 604.0 103°58′10.3″ 31°08′14.4″ 2017.05 CX14 Pengzhou, Sichuan 604.0 103°59′53.1″ 31°07′56.7″ 2017.05 CX15 Pengzhou, Sichuan 599.0 103°59′38.7″ 31°06′40.3″ 2017.05 CX16 Pengzhou, Sichuan 590.0 103°59′23.3″ 31°03′08.5″ 2017.05 CX17 Pengzhou, Sichuan 568.0 104°00′57.8″ 31°01′03.1″ 2017.05 CX18 Xindu, Sichuan 542.0 103°59′46.8″ 30°53′59.6″ 2017.05 CX19 Dujiangyan, 626.0 103°39′14.5″ 30°49′48.5″ 2017.05 Sichuan CX20 Dujiangyan, 592.0 103°40′21.5″ 30°46′38.8″ 2017.05 Sichuan CX21 Dujiangyan, 626.0 103°40′45.8″ 30°50′45.8″ 2017.05 Sichuan CX22 Chongzhou, 573.0 103°39′58.3″ 30°43′46.5″ 2017.05 Sichuan CX23 Fuqing, Fujian 122.0 119°23′15.2″ 25°44′28.1″ 2018.05 CX24 Zhenba, Shanxi 657.0 108°01′39.5″ 32°32′45.6″ 2018.06 Trop J Pharm Res, April 2021; 20(4): 758 Chen et al Epinephrine hydrochloride injection was injection of 0.1 % epinephrine hydrochloride, rats purchased from Grand Pharmaceuticals (Beijing, in all groups except for the control group were China), and 0.9% sodium chloride injection was placed in an iced water bath for 5 min. Thirty obtained from Sichuan Kelun Pharmaceuticals minutes after gavage on day-8, rats in all groups (Sichuan, China). Compound danshen tablets were anesthetized with 10% chloral hydrate, and were obtained from Yunnan Baiyao Group Co., blood was collected from the abdominal aorta. Ltd. (Yunnan, China). Treatment and assessment of blood samples Specific pathogen-free male and female Sprague–Dawley rats (180 – 220 g) were First, parameters from whole-blood samples purchased from Chengdu Dashuo Experiment were measured by an automatic Animals (Chengdu, China). All rats were fed in a hemorrheometer (SA-6000; Succeeder). Then, temperature-controlled facility with relative samples were centrifuged (3000 rpm for 30 min humidity of 65% and a 12-h light–dark cycle. at room temperature), the supernatant separated, and analyzed. Blood-activating effect of CX extracted using different solvents Determination of spectrum–effect relation- ship Sample preparation Sample preparation CX samples (250 g/batch) were ground into powder, soaked with 70% ethanol for 10 h, and The method of sample preparation was identical extracted eight times with 70% ethanol by to that described as sample preparation in blood- seepage. The extracts were combined, and activating effect of CX extracted using different concentrated to a flow extract (30 mL) under solvents. reduced pressure, then dispersed with the appropriate amount of pure water. Then, they Grouping and treatment of rats were extracted thrice with petroleum ether, dichloromethane, ethyl acetate or N-butanol, Rats were divided randomly into 27 groups: respectively. Extracts of each part were blank; model; CDT; CX1–CX24. Rats in the combined and concentrated under reduced CX1–CX24 groups were given the corresponding pressure, and diluted to 250 mL with 0.5% extracts at 1 g/kg/day (bodyweight) for 7 days, carboxymethylcellulose sodium (CMC-Na) to respectively. Other treatments were identical to obtain samples. those described as grouping and treatment of rats in blood-activating effect of CX extracted Grouping and treatment of rats using different solvents.
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