Determination of 13 Trace Elements in Anshun Thorn Pear (Rosa roxburghii) and Golden Thorn Pear (Rosa sterilis S. D. Shi) by ICPMS

Wei CHEN Kaibin LI Qiuyue KONG Sibu MA Wen YANG

Abstract [Objectives]This study was conducted to establish an analytical method for the establishment of 13 trace elements in Anshun thorn pear （Rosa roxburghii） and golden thorn pear （Rosa sterilis S. D. Shi）.[Methods]Nitric acid was used as the digestion system for microwave digestion. Inductively coupled plasma mass spectrometry （ICPMS） was used to determine the trace elements in thorn pear and golden thorn pear.[Results]Be， Ti， V， Cr， Mn， Co， Ni， As， Sr， Mo， Cd， Sb and Hg were determined by ICPMS method in thorn pear and golden thorn pear. The correlation coefficients of the regression equations of the elements were higher than 0.996; the precision RSD was lower than 2.87%; and the recovery ranged from 94.3% to 109.8%.[Conclusions]The method has the advantages of high sensitivity， good accuracy and simultaneous detection of multiple elements， and provides a reference method and theoretical basis for determining trace elements in thorn pear and golden thorn pear and other fruits.

Key words Inductively coupled plasma mass spectrometry; Microwave digestion method; Rosa roxburghii; Rosa sterilis S. D. Shi; Element analysis

Thorn pear （Rosa roxburghii）， also known as Saosihua and Songchungui， refers to the fruit of a perennial deciduous shrub in Rosaceae[1]， mainly distributed in Guizhou， Sichuan， Yunnan and other places in China， especially the most abundant in Guizhou[2]. R. roxburghii has a high edible value and medicinal value， and is recorded in "Guizhou Traditional Chinese Medicinal Material and Ethnic Minority Medicine Quality Standard" （2003 Edition）[3]. In recent years， studies have shown that the ingredients contained in thorn pear have antioxidation， immunityimproving， anticancer and antiaging effects[4-5]. Golden thorn pear is the fruit of R. sterilis S. D. Shi which was published in 1985 by Mr Shi from Guizhou Botanical Garden and is a special plant in Guizhou Province[6]. The fruit of golden thorn pear is golden yellow， and is orangeyellow when it is mature， and thorns on the surface of the fruit would fall off when it is fully mature. The flesh of golden thorn pear is thicker than ordinary thorn pear， orangeyellow， very crisp， sweet and refreshing， and moderately sour （hence the name）. Anshun golden thorn pear is authorized by the Protection of New Varieties of Plants and is the first authorized new plant variety obtained in Guizhou Province.

The determination of trace elements plays an important role in the analysis of thorn pear and golden thorn pear as they play an important role in physiological and biochemical reactions and metabolism. At present， the research on the fruits of thorn pear and golden thorn pear is mainly carried out in vitamins， polyphenols， flavonoids， triterpenoids and amino acids[7-8]， and few studies have been conducted on trace elements， mainly applying atomic absorption spectrometry （AAS） and inductively coupled plasma optical emission spectrometry （ICPOES）[9-11]. Inductively coupled plasma mass spectrometry （ICPMS） has the advantages of low detection limit， wide linear range， low interference， and simultaneous determination of multiple elements with a very low concentration， and has been widely used in the analysis of trace elements in foods and medicines[12-13]. In this study， microwave digestionICPMS method was used to determine the trace elements in the fruits of thorn pear and golden thorn pear， laying a foundation for the research， quality control and deep processing of Guizhou thorn pear and golden thorn pear.

Materials and Methods

Materials and reagents

The samples were picked from Anshun City， Guizhou Province， and the collected samples were heavier than 1 kg.

Reagents： Wahaha pure water （Guiyang Wahaha Beverage Co.， Ltd.）， nitric acid （guarantee reagent， Sinopharm Group）， Cd standard solution （GSB0417122004）， Hg standard solution （GSB0417292004）， and As standard solution （GSB0417142004）， all of which were purchased from National Center of Analysis and Testing for Nonferrous Metals and Electronic Materials; Be， Ti， V， Cr， Mn， Co， Ni， Sr， Mo and Sb multielement mixed standard solution （N9301721， PerkinElmer）.

Instruments and equipment

NexION2000 inductively coupled plasma mass spectrometer （PerkinElmer， USA）; ETHOS UP microwave digestion instrument （Beijing LabTech Instruments Co.， Ltd.）; an electronic analytical balance （0.001 g， Shimadzu Corporation）.

Methods and Results

Sample pretreatment

The sample collected was taken back to the laboratory and rinsed with tap water and purified water. It was placed in a blast drying oven at 60 and dried to constant weight， and then pulverized for use.

A certain amount of the sample （accurate to 0.000 1 g） was accurately weighed into the special PTFE digestion tank of the microwave digestion device， added with 10 ml of concentrated nitric acid， followed by standing overnight to allow digestion. The digestion program is shown in Fig. 1. After the digestion was completed， the solution was subjected to acid removal on an acid meter and concentrated to 1-2 ml. The solution was transferred to a 50 ml graduated cylinder， and diluted to constant volume with purified water， for later detection.

Preparation of standard solutions

An appropriate amount of the mercury standard stock solution was precisely measured and diluted with 10% nitric acid solution to prepare 0， 10， 20， 40 and 80 ng/ml Hg standard solutions， respectively. The solutions were prepared freshly for use.

An appropriate amount of the 10 g/ml mixed standard solution （Be， Ti， V， Cr， Mn， Co， Ni， Sr， Mo and Sb） supplied by PerkinElmer was diluted to 0， 10， 20， 40 and 80 ng/ml solutions.

Working parameters of ICPMS

The working parameters of ICPMS are shown in Table 1. The sample was flushed for 35 s; the determination was delayed by 15 s; the data collection adopted the fully quantitative mode; eachdetermination was repeated for 3 times.

Methodology validation

Linear regression equation and correlation coefficient

The linear regression equation and correlation coefficient of each element are shown in Table 2. The correlation coefficients were all larger than 0. 996， and the linear relationship was good.

Precision and reproducibility

The reference solution with the middle concentration was continuously injected for 6 times， obtaining the response value CPS， which was the precision， the RSD range of which was 1.23%-2.87%， indicating that the precision of the instrument is good. Six parts of parallel samples were prepared and injected， and the average content of each element was obtained as the result， which had a RSD range of 0.96%-4.92%. The results are shown in Table 2.

Detection limit

A blank solution was prepared referring to the method under "Sample pretreatment"， except that no sample was added （other operations were the same as the preparation of the test solution）. The blank sample was measured continuously for 11 times. The standard deviation of 3 times of the response value was divided by the slope of the corresponding element standard curve， obtaining the detection limit of corresponding element[14]， as shown in Table 2.

Recovery test

Various elements in the sample were subjected to a recovery test. A certain amount of the sample was accurately weighed （0.3 g， accurate to 0.000 1 g）， and the mixed standard solution was added. Under the same conditions， the microwave digestion treatment was carried out and the recovery of each element was obtained， as shown in Table 3. The recovery ranged from 94.3% to 109.8%. The method is accurate and reliable.

Sample determination results

The thorn pear and golden thorn pear samples were prepared according to "Sample pretreatment"， and the contents of trace elements contained in the samples were determined by ICPMS under the optimal working conditions， as shown in Table 3.

Results and Discussion

Studies have shown that 50% to 70% of enzymes in the human body require the participation of metal elements or as an enzyme activator. In the case of excessive intake of heavy metals， the activity of the biological enzyme is inhibited and destroyed. The heavy metals entering the human body mainly combine with organic components in the body to form metal complexes or metal chelate compounds which change the conformation of the active sites by binding to the inactive sites of enzymes， so that the activity of the enzymes is weakened or even lost， thereby showing toxicity[15]. In this study， heavy metal elements such as As， Cd and Hg were also measured， referring to the limits in current "Green Trade Standards of Importing & Exporting Medicinal plants & Preparations"， "Chinese Pharmacopoeia" （2015 Edition）， GB1488294 "Limited Concentrations of Radioactive Materials in Foods"： Cd  0.3 mg/kg， Hg  0. 2 mg/kg， As  2. 0 mg/kg[12]. As， Cd， and Hg were all detected in the above samples tested， but did not exceed the limits.

In this study， samples was pretreated by microwave digestion， and Be， Ti， V， Cr， Mn， Co， Ni， As， Sr， Mo， Cd， Sb and Hg were determined by ICPMS method in thorn pear and golden thorn pear. The samples were completely digested， and the results were accurate and reliable. The correlation coefficients of the regression equations of the elements were higher than 0.996; the precision RSD was lower than 2.87%; and the recovery ranged from 94.3% to 109.8%. The method has the advantages of high sensitivity， good accuracy and simultaneous detection of multiple elements， and provides a reference method and theoretical basis for determining trace elements in thorn pear and golden thorn pear and other fruits.

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