Evaluation of Wild Paeonia veitchii Lynch from Sichuan Based on Moisture, Ash and Extract Contents

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1. College of Pharmacy, Southwest Minzu University, Chengdu 610041, China; 2. Ethnic Medicine Institute, Southwest Minzu University, Chengdu 610041, China

Abstract [Objectives] The research aimed to determine the contents of moisture, ash and extract of wild Paeonia veitchii Lynch produced in Sichuan, so as to provide the basis for evaluating quality of wild P. veitchii Lynch from Sichuan. [Methods] Moisture, ash and extract contents of wild P. veitchii Lynch were respectively determined according to the fourth part of the Chinese Pharmacopoeia of 2015 version, and correlation analysis and TOPSIS analysis were used for data processing. [Results] 12 batches of samples all corresponded with the provision that moisture of P. veitchii Lynch should not exceed 14.0% in the Standard of Traditional Chinese Medicine in Sichuan Province of 2010 version; 11 batches of samples all corresponded with the provisions that total ash and acid insoluble ash of P. veitchii Lynch should not exceed 7.0% and 1.0% in the Standard of Traditional Chinese Medicine in Sichuan Province of 2010 version; average content of alcohol-soluble extract by hot dipping from 12 batches of samples was 42.29%, which was not less than 30.0%. In P. veitchii Lynch, moisture and acid insoluble ash showed positive correlation with altitude, while total ash and extract content were negatively correlated with altitude. Taking moisture, ash and extract as evaluation indexes, No.9 P. veitchii Lynch sample was the best. [Conclusions] Moisture and ash of P. veitchii Lynch corresponded with the provisions of the Chinese Pharmacopoeia of 2015 version and the Standard of Traditional Chinese Medicine in Sichuan Province of 2010 version, while extract content was higher than provision of the standard. The research could provide reference basis for rational utilization and quality evaluation of P. veitchii Lynch resources.

Key words Paeonia veitchii Lynch, Moisture, Ash, Extract, Content determination

1 Introduction

Radix Paeoniae Rubra is dry root fromPaeoniaveitchiiLynch, and it is one of sources of Paeoniaeradix Rubra in theChinesePharmacopoeiaof 2015 version[1]. In theStandardofTraditionalChineseMedicineinSichuanProvinceof 2010 version[2], it is recorded that Radix Paeoniae Rubra is dry root and rhizome ofP.veitchiiLynch var.Woodwardii(Stapf. ex Cox.) Stern,P.veitchiiLynch var.unifloraK. Y. Pan,P.maireiLévl.,P.obovateMaxim., andP.obovateMaxim. var.Willmottiae(Stapf.) Stern. It has the functions of clearing heat, cooling blood, removing blood stasis and relieving pain, and is mainly used to treat heat toxin with skin eruption, hematemesis and epistaxis, redness and swelling of eyes, liver depression and hypochondriac pain, injury of fall and flutter, welling-abscess swellings and sores.

P.veitchiiLynch is traditional medicinal material commonly used in China, and has a long history of application. It is mainly produced in Sichuan, and is a kind of local medicine.P.veitchiiLynch is widely distributed in Qinghai-Tibet Plateau of northwest Sichuan, with the altitude of 3 000-3 500 m. Via the research on chemical composition ofP.veitchiiLynch[3-8], it is found thatP.veitchiiLynch mainly contains monoterpenes and their glycosides: paeoniflorin, oxypaeoniflorin, benzoylpaeoniflorin, paeoniflorin, benzoyl hydroxy paeoniflorin; flavonoids and their glycosides: catechin, kaempferol, kaempferol-3,7-diglucoside; tannins: gallic acid, 1,2,3,4,6-pentagallic glucose; aromatic acids and their esters: benzoic acid, salicylic acid, palmitic acid, ethyl gallate; phenols and their glycosides: paeonol, paeonol glycoside; amino acids, volatile oil,etc. Pharmacological experiments[9-12]prove thatP.veitchiiLynch has the functions of antithrombotic, protecting heart, antitumor, scavenging active oxygen free radical, and lowering blood sugar. In this paper, contents of moisture, ash and extract of wildP.veitchiiLynch from Sichuan were determined, which aimed to provide test basis for evaluating quality of medicinal materials of wildP.veitchiiLynch from Sichuan.

2 Materials and methods

2.1MaterialsMETTLER AE240 electronic analytical balance (Shanghai METTLER-TOLEDO Instrument Co., Ltd.), DHG-9240A type of electric constant-temperature blast drying oven (Shanghai Jinghong Experimental Equipment Co., Ltd.), SX2 box-shape resistance furnace(Changsha Yuandong Electric Furnace Factory), HH-2 type of digital-display constant-temperature water bath(Guohua Electric Co., Ltd.). Ethanol, dilute hydrochloric acid and other reagents were all AR.

P.veitchiiLynch was collected from Aba Prefecture and Ganzi Prefecture of Sichuan Province during 2017-2018, and sources of each samples were shown as Table 1. Via identification by Professor Liu Yuan from Ethnic Medicine Institute of Southwest Minzu University, they wereP.veitchiiLynch.

Table1CollectioninformationofPaeoniaveitchiiLynch

No.Collection site Time Altitude∥mS1Zhonggu Village, Yala Township, Kangding City, Ganzi Prefecture2017-07-103 160S2Lengleng Village, Geka Township, Daofu County, Ganzi Prefecture2017-07-113 320S3Shawan Village, Geka Township, Daofu County, Ganzi Prefecture2017-07-113 310S4Jiapa Village, Daofu County, Ganzi Prefecture2017-07-113 445S5Gerenda Village, Chonggu Township, Ganzi County, Ganzi Prefecture2017-07-133 540S6Ridu Village, Ganzi County, Ganzi Prefecture2017-07-133 610S7Tatuo Village, Ganzi County, Ganzi Prefecture2017-07-133 530S8Gage Village, Ganzi County, Ganzi Prefecture2017-07-133 560S9Yiniang Village, Luhuo County, Ganzi Prefecture2017-07-123 240S10Dari Village, Gangmuda Township, Rangtang County, Aba Prefecture2018-07-073 308S11Xiaoyili Village, Puxi Township, Rangtang County, Aba Prefecture2018-07-102 961S12Balang Mountain, Gengda Town, Wenchuan County, Aba Prefecture2018-05-234 432

2.2Methods

2.2.1Moisture determination. According to the second moisture determination method (drying method) in the fourth part of theChinesePharmacopoeiaof 2015 version[13], 2.0 g ofP.veitchiiLynch powder (sieved through No.3 sieve) was weighed and laid flat in a flat weighing bottle that has been dried to constant weight, and its thickness did not exceed 5 mm. The weight was weighed, and bottle cap was open, and it was dried for 5 h in 105 ℃ of oven. After covering the bottle cap, it was taken out and set in dryer to cool for 30 min. It was taken out for precise weighing, and then it was dried for 1 h in above temperature. After cooling, it was weighed. Above steps were repeated until that the two consecutive weight difference was within 5 mg. According to weight loss of medicinal materials, moisture of medicinal materials was calculated.

2.2.2Ash determination. (i) Determination of total ash. According to the determination method of total ash in the fourth part of theChinesePharmacopoeiaof 2015 version[13], 3.0 g ofP.veitchiiLynch powder (sieved through No.3 sieve) was weighed and set in crucible burning to constant weight. It was weighed precisely and heated slowly, and the temperature gradually increased to 500-600 ℃ when it was fully carbonized, making it completely ashed to constant weight. According to residue weight, content of total ash in sample was calculated.(ii) Determination of acid insoluble ash. According to the determination method of acid insoluble ash in the fourth part of theChinesePharmacopoeiaof 2015 version[13], ash obtained in Section2.2.1was taken, and 10 mL of dilute hydrochloric acid was added into the crucible carefully, and the crucible was covered with a glass-surface vessel. After that, it was set in water bath to heat for 10 min. Glass-surface vessel was washed by 5 mL of hot water, and washing solution was merged into crucible. It was filtered by ashless filter paper, and the residue in the crucible was washed on the filter paper with water until the solution did not show chloride reaction. The filter residue and filter paper were moved into the same crucible, and it was dried in above temperature to constant weight. According to residue weight, content of acid insoluble ash in sample was calculated.

2.2.3Extract determination. (i) Comparison on the determination methods of extract. It was measured according to extract determination method in the fourth part of theChinesePharmacopoeiaof 2015 version[13].

Cold dipping method: 4 g of test sample was taken in 250 mL of conical flask, and 100 mL of solvent was added precisely. Then it was pluged, soaked and quickly filtered with dry filter after 6 h of shaking and 18 h of standing. 20 mL of continuous filtrate was taken precisely to set in evaporating dish dried to constant weight. After dried in a water bath, it was dried for 3 h at 105 ℃ and then cooled for 30 min in dryer. After that, it was quickly weighed. The dried product was used to calculate the content of extract in the test sample, and results were shown in Table 2.

Hot dipping method: 2 g of test sample was taken in 100 mL of conical flask, and 50 mL of solvent was added precisely. Then it was pluged and weighed. After 1 h of standing, it was heated to boiling, and maintained slight boiling for 1 h. After cooling, conical flask was taken, pluged and weighed again. The lost weight was made up, and then it was shaken evenly. Dry filter was used to filtrate, and 25 mL of filtrate was precisely taken to set in evaporating dish dried to constant weight. After steaming in a water bath, it was dried for 3 h at 105 ℃ and then cooled for 30 min in dryer, and its weight was quickly weighed. The dried product was used to calculate the content of extract, and results were shown in Table 2.

Table2Determinationresultsofcoolandhotdippingmethodsbydifferentsolvents

%

Test results showed that hot dipping method of 50% ethanol could obtain higher extract content, so hot dipping method of 50% ethanol was used to determine extract content of sample.

(ii) Content determination of alcohol-soluble extract. According to hot dipping method in the fourth part of theChinesePharmacopoeiaof 2015 version[13], ethanol concentration was 50%, and extract in sample was calculated.

3 Results and analyses

Determination results of moisture, ash and extract contents from 12 batches ofP.veitchiiLynch were shown in Table 3.

No.Moisture Total ashAcid insoluble ash Extract S17.88±0.045.68±0.040.58±0.0140.51±0.69S27.30±0.117.27±0.151.43±0.0149.13±0.27S36.20±0.026.54±0.060.72±0.0245.95±0.81S47.86±0.086.49±0.060.88±0.0246.63±0.65S57.98±0.106.15±0.080.94±0.0149.16±0.94S68.10±0.056.51±0.120.97±0.0147.75±0.78S78.33±0.026.53±0.060.96±0.0246.59±0.63S87.82±0.065.32±0.040.73±0.0049.12±0.59S98.29±0.034.39±0.030.58±0.0146.66±0.64S108.33±0.025.78±0.040.83±0.0141.92±0.51S118.42±0.165.27±0.060.73±0.0135.71±0.48S128.30±0.035.00±0.080.76±0.0134.33±0.48

Seen from Table 3, the highest moisture content of 12 batches ofP.veitchiiLynch was 8.42%, the minimum was 6.20%, and mean moisture content was 7.90%. 12 batches of samples all corresponded with the provision that moisture ofP.veitchiiLynch should not exceed 14.0% in theStandardofTraditionalChineseMedicineinSichuanProvinceof 2010 version[2]. The highest total ash content of 12 batches ofP.veitchiiLynch was 7.27%, the minimum was 4.39%, and mean total ash content was 5.91%. Among them, 11 batches of medicinal materials corresponded with the provision that total ash content ofP.veitchiiLynch should not exceed 7.0% in theStandardofTraditionalChineseMedicineinSichuanProvinceof 2010 version[2]. The highest content of acid insoluble ash of 12 batches ofP.veitchiiLynch was 1.43%, the minimum was 0.58%, and mean acid insoluble ash content was 0.84%. Among them, 11 batches of medicinal materials all corresponded with the provision that acid insoluble ash ofP.veitchiiLynch should not exceed 1.0% in theStandardofTraditionalChineseMedicineinSichuanProvinceof 2010 version[2]. The highest extract content of 12 batches ofP.veitchiiLynch was 49.16%, the minimum was 34.33%, and mean extract content was 44.46%.

3.1CorrelationanalysisCorrelation analysis results between moisture, ash, extract contents ofP.veitchiiLynch and altitude were shown in Table 4. Seen from Table 4, moisture and acid insoluble ash ofP.veitchiiLynch showed positive correlation with altitude, while total ash and extract contents were negatively correlated with altitude.

3.2TOPSISanalysisCoefficient of variation method was used to calculate the weights of moisture, ash, extract contents ofP.veitchiiLynch[14], and results were shown in Table 5. According to weight of each component, TOPSIS analysis of the standardized data was conducted by DPS software with moisture, total ash, and acid insoluble ash in low-quality indexes. The results in Table 6 showed that the quality of No.9P.veitchiiLynch was the best by taking moisture, ash and extract as evaluation indexes.

Table4Correlationbetweenmoisture,ash,extractcontentsofPaeoniaveitchiiLynchandaltitude

Altitude Moisture TotalashAcidinsolubleashExtractAltitude Moisture 0.154Total ash-0.105-0.500Acid insoluble ash0.081-0.1580.776∗∗Extract -0.186-0.3560.4930.429

Table5Weightresultsofmoisture,ash,extractcontentsofPaeoniaveitchiiLynchbycoefficientofvariationmethod

Item Moisture TotalashAcidinsolubleashExtract Coefficient of variation0.078 7 0.138 3 0.270 5 0.116 3 Weight0.130 4 0.229 0 0.448 0 0.192 6

Table6TOPSISevaluationresultsofmoisture,ash,extractcontentsofPaeoniaveitchiiLynch

Sample Index CIRanking Sample Index CIRanking S10.809 42S70.336 110S20.149 812S80.644 93S30.619 74S90.898 61S40.415 28S100.478 67S50.370 69S110.616 45S60.332 211S120.581 46

4 Conclusions and discussions

Taking determination results into consideration, except total ash and acid insoluble ash in S2 sample exceeded the provision that total ash and acid insoluble ash ofP.veitchiiLynch should not exceed 7.0% and 1.0% in theStandardofTraditionalChineseMedicineinSichuanProvinceof 2010 version, other samples all corresponded with the requirements. Meanwhile, there was larger difference between the maxima and the minima of moisture, ash and extract contents from 12 batches of wildP.veitchiiLynch in different sites of Sichuan, which illustrated that there were differences in the quality of wildP.veitchiiLynch from different sites of Sichuan.

There is no determination requirement on extract ofP.veitchiiLynch in theStandardofTraditionalChineseMedicineinSichuanProvinceof 2010 version. Main components ofP.veitchiiLynch are glycosides, which are easy to dissolve in water, alcohol and other solvents. Meanwhile, pre-test results of extract showed that extract content by 50% ethanol was the highest, so 50% ethanol was selected as solvent to determine extract content. The determination results showed that there was larger difference in extract content ofP.veitchiiLynch from different sites, and whole extract content of wildP.veitchiiLynch collected from Ganzi County of Ganzi Prefecture was overall higher, while extract content ofP.veitchiiLynch from Balang Mountain, Gengda Town, Wenchuan County, Aba Prefecture was the lowest. Maybe the differences in the environmental and climatic conditions of plant growth lead to the difference in the extract content of medicinal materials. Based on test results, it is suggested that extract content ofP.veitchiiLynch should not be less than 30%. The research could provide basis for further establishing quality standard ofP.veitchiiLynch.