Determination and Multidimensional Analysis of 29 Inorganic Elements in Cardamine tangutorum O. E. Schulz of Different Origins

Xiaoxia CAI, Bochao LI, Junzhang QUBIE, Yuebu HAILAI, Jianlong LAN, Wenbing LI, Yuan LIU

1. College of Pharmacy, Southwest Minzu University, Chengdu 610041, China; 2. Sichuan Qiang Yi Medicinal Resources Protection and Utilization Technology Engineering Laboratory, Chengdu 610041, China; 3. Institute of Qinghai-Tibetan Plateau Research, Southwest Minzu University, Chengdu 610041, China; 4. Ethnic Medicine Institute, Southwest Minzu University, Chengdu 610041, China

Abstract [Objectives] This study aimed to determine the contents of inorganic elements in Cardamine tangutorum O. E. Schulz samples from different origins to provide a scientific reference for the quality control, safety evaluation and clinical medicinal use of C. tangutorum O. E. Schulz. [Methods] The contents of Al, As, B, Ba, Ca, Cd, Co, Cr, Cu, Fe, Hg, K, Mg, Mn, Mo, Na, Ni, P, Pb, Pd, Rb, Se, Si, Sn, Sr, Ti, Tl, V and Zn in the C. tangutorum O. E. Schulz samples were determined simultaneously by wet digestion and inductively coupled plasma optical emission spectroscopy (ICP-OES), and the determination results were analyzed by principal component analysis. [Results] A total of 28 inorganic elements were detected in the samples of C. tangutorum O. E. Schulz from different origins. The contents of heavy metals did not exceed the limits. Among the elements, the determined values of Hg in all samples were below the detection limit, so it was not detected. The contents of Ca, Al, P and Fe were relatively high, and they were essential nutrients in the human body. There were significant differences among different elements. Samples from different origins show similar patterns in the distribution of main elements. Correlation analysis shows that 15 pairs of inorganic elements out of 24 elements had extremely significant positive correlations, and 26 pairs of elements had significant positive correlations. The principal component analysis determined Al, As, Ba, Ca, Cd, Fe, Mn , Ni, P, Pb, Pd, Se, V and Zn were the characteristic inorganic elements in the samples of C. tangutorum O. E. Schulz. [Conclusions] The contents of toxic heavy metals in C. tangutorum O. E. Schulz samples from different origins met the edible requirements. The method was simple, rapid and accurate, and could be used to analyze and determine the inorganic elements in the samples of C. tangutorum O. E. Schulz, providing a theoretical basis for the better development and utilization of C. tangutorum O. E. Schulz.

Key words Cardamine tangutorum O. E. Schulz, Inorganic element, Inductively coupled plasma optical emission spectroscopy (ICP-OES), Principal component analysis

1 Introduction

Medicinal and edible plants were gained attention and recognition worldwide. Micro-inorganic elements were one of the material bases for the efficacy of medicinal and edible plants. Trace elements participate in the physiological reactions of hormones, vitamins and enzymes in organisms. Some essential inorganic elements also were special physiological functions in many macromolecules. Therefore, the research on the trace elements contained in medicinal and edible plants was an important part of the quality evaluation of medicinal and edible plants; and the safety evaluation of inorganic elements contained in medicinal and edible plants has become the research direction of bioinorganic chemistry, traditional Chinese medicine pharmacology and toxicology[1].

CardaminetangutorumO. E. Schulz (Cruciferae) was usually used as medicine with its dried whole herbs, also known as Shigecai and Shijiecai. The young stems and leaves ofC.tangutorumO. E. Schulz were a popular vegetable for the Tibetan, Qiang and Yi peoples who live on the southeastern edge of the Qinghai-Tibet Plateau, with a good taste. In Tibetan Medicine, it was used for treating yellow-water sores and arthralgia and myalgia[2-4]. Cardamine spp. contained flavonoids, volatile oils, nitrogen-containing compounds, and so on[5-8]. Flavonoids was antioxidant, anti-inflammatory, antibacterial, anticancer and antiviral effects[9-11].

There were no reports about the quality control and evaluation ofC.tangutorumO. E. Schulz from the perspective of beneficial trace elements and heavy metals. In this study, the contents of 29 kinds of inorganic elements in different batches ofC.tangutorumO. E. Schulz samples by wet digestion and inductively coupled plasma optical emission spectroscopy (ICP-OES), so as to provide a reference for the safety assessment and resource development ofC.tangutorumO. E. Schulz.

2 Materials and methods

2.1 MaterialsThe main instruments and equipment used were inductively coupled plasma emission optical emission spectroscopy (ICP-OES) (6300 Radial, Thermo, America) and dual-range analytical balance[METTLER AE240, Mettler-Toledo Instruments (Shanghai) Co., Ltd.]. The main reagents and drugs used included Aluminum (Al), arsenic (As), boron (B), barium (Ba), calcium (Ca), cadmium (Cd), cobalt (Co), chromium (Cr), copper (Cu), iron (Fe), mercury (Hg), potassium (K), magnesium (Mg), manganese (Mn), molybdenum (Mo), sodium (Na), nickel (Ni), phosphorus (P), lead (Pb), palladium (Pd), rubidium (Rb), selenium (Se), silicon (Si), tin (Sn), strontium (Sr), titanium (Ti), thallium (Tl), vanadium (V) and zinc (Zn) standard solutions (National Research Center for Certified Reference Materials, 1 mg/mL), water (ultrapure), nitric acid (guaranteed reagent) and perchloric acid (guaranteed reagent).

The samples ofC.tangutorumO. E. Schulz (Table 1) were collected in different areas of Aba Tibetan and Qiang Autonomous Prefecture in Sichuan Province. They were identified by Professor Liu Yuan of Southwest Minzu University. The samples were pulverized and passed through No.4 sieve for later use.

2.2 Methods

2.2.1Preparation of test solution. A certain mount (1.0 g) of the powder of eachC.tangutorumO. E. Schulz sample was soaked in 8 mL of nitric acid for about 30 min, added with 2 mL of perchloric acid, and let standard at 105 ℃. The above operation was repeated until the liquid evaporated and the sample power turned white. The digestion solution was filtered and diluted to 10 mL in a colorimetric tube as the test solution. The blank sample was prepared into a blank control solution in the same way. Three duplicates were arranged for each sample.

2.2.2Sample determination. The conditions of ICP-OES were as follows: RF power, 1.15 kW; plasma gas flow rate, 1.5 L/min; auxiliary gas flow rate, 0.5 L/min; atomizer flow rate, 0.55 L/min; pump speed, 50 rpm; observation height, 10 mm; reading time, 5 s; determination times, 3; stabilization delay, 15 s. Under the set working conditions, the mixed standard and sample solution were determined.

2.2.3Data analysis. SPSS 21.0 statistical software was used to perform correlation analysis and principal component analysis on the data.

3 Results and analysis

3.1 Element determinationThe contents of inorganic elements in theC.tangutorumO. E. Schulz samples were shown in Table 2.

As shown in Table 2,C.tangutorumO. E. Schulz was rich in Ca, Fe, Al, P and Pd. There were great differences in the content of different inorganic element among the samples ofC.tangutorumO. E. Schulz from different origins. The content of Ca in from different producing areas was significantly higher than those of other elements.C.tangutorumO. E. Schulz from Meiwo Township, Xiaojin County, Aba Prefecture (A2) showed the highest Ca content, reaching 618.3 μg/g, and that from Chuanzhusi, Aba Prefecture, Sichuan Province (A5) showed the lowest Ca content, only 276.8 μg/g. The content of Sn in theC.tangutorumO. E. Schulz samples from different origins was very low, and the highest was only 0.010 6 μg/g (A1, Meiwo Township, Xiaojin County, Aba Prefecture). At the same time,C.tangutorumO. E. Schulz also contained Mo, Ni, Si, Sr, V, Cu, Cd, B and other essential trace elements. Hg and Cr elements were not detected in all the samples. Although some toxic heavy metals such as Pb and Cd were detected, the contents were very low.

Table 2 Contents of inorganic elements in Cardamine tangutorum O. E. Schulz samples from different origins (μg/g, n=3)

3.2 Analysis of the inorganic element spectrumBased on the determination results and referring to the analysis methods of wild ginseng[13]and fleece-flower root[14], 24 kinds of inorganic elements (Al, As, B, Ba, Ca, Cd, Co, Cr, Cu, Fe, Mn, Mo, Na, Ni, P, Pb, Pd, Se, Si, Sn, Sr, Ti, V, Zn) were screened to draw the element content distribution map (Fig.1). As shown in Fig.1, although there were great differences in the contents of the 24 kinds of inorganic elements among the 6 producing areas, the overall trend was the same. The contents of Ca, Al, P and Fe were significantly higher than those of other elements, which could provide reference for the quality evaluation ofC.tangutorumO. E. Schulz.

3.3 Correlation analysisBased on the results above, correlation analysis was carried out on the 24 inorganic elements inC.tangutorumO. E. Schulz with SPSS 21.0 software. As shown in Table 3, there were certain correlations among the inorganic elements inC.tangutorumO. E. Schulz. Among them, 15 pairs of elements had extremely significant positive correlations (**P<0.01), and 26 pairs of elements had significant positive correlations (*P<0.05). These positive correlations indicated that there was a synergy between these elements,C.tangutorumO. E. Schulz may absorb these elements in a certain proportion, and there was certain selectivity in the absorption of these elements. There was no correlation between the Si element content distribution and other elements.

Fig.1 Distribution curve of inorganic elements in Cardamine tangutorum O. E. Schulz samples from different origins

Table 3 Correlations among the 24 kinds of inorganic elements in Cardamine tangutorum O. E. Schulz

3.4 Principal component analysisPrincipal component analysis was widely used in the analysis and research of inorganic elements in Chinese herbal medicines[15-17]. In this study, the contents of 24 inorganic elements determined in theC.tangutorumO. E. Schulz samples were subjected to principal component analysis after standardization treatment using SPSS 21.0 software. As shown in Table 4, the cumulative variance contribution rate of the first four principal components reached 97.88% (>85%[18]), so they were selected as they represented almost all the information about the inorganic elements in theC.tangutorumO. E. Schulz samples. In order to better investigate the influence of each element on different principal components, the maximum variance method was used to perform rotation analysis on the initial load matrix to obtain the factor loading matrix. As shown in Table 5, the coefficients of As, Ba, Cd, Fe, Mn, Ni, Pb and Se in the primary principal components were relatively large, and they were the feature vector that had the greatest impact on the primary principal components. The elements that had the greatest influence on the secondary principal components were Al, Ca, P, Pd, V and Zn. The elements that had the greatest influence on the tertiary principal components were Na, Si and Sn. The element that had the greatest influence on the quaternary principal components was Ti. As 59.09% of the total variance came from the first two principal components, it could be considered that Al, As, Ba, Ca, Cd, Fe, Mn, Ni, P, Pd, Pb, Se, V and Zn were the characteristic inorganic elements in the samples ofC.tangutorumO. E. Schulz.

Table 4 Eigenvalues and contribution rates of principal components

Table 5 Factor loading matrix after rotation transformation

4 Discussions

In this experiment, it was found that the contents of Ba, Ca, Pd and Se in theC.tangutorumO. E. Schulz samples from different origins were significantly higher than those of other inorganic elements, and they were important inorganic elements in the growth ofC.tangutorumO. E. Schulz. There were no limit requirements for heavy metals under the category ofC.tangutorumO. E. Schulz inDictionaryofChineseEthnicMedicine. Based on the important role of heavy metals in the safety of medicinal and edible plants, this article refered to the guidance values of the consistency limit of heavy metals and harmful elements inChinesePharmacopoeia(2020 Edition, General Principle 9302): Pb (≤ 5.0 mg/kg), Cd (≤ 1 mg/kg), As (≤ 2 mg/kg), Hg (≤ 0.2 mg/kg), Cu (≤ 20.0 mg/kg). The contents of 29 inorganic elements in the samples ofC.tangutorumO. E. Schulz from different origins were determined. None of them exceeded the limits, and the heavy metal Hg was not detected in all the samples. It was showed that the contents of toxic heavy metals in the samples ofC.tangutorumO. E. Schulz from different origins met the food requirements. The contents of Pb, Cd, Hg and other harmful elements in the samples ofC.tangutorumO. E. Schulz were low, and those of Ca, Al, Fe, Zn, P and other beneficial elements were relatively high, in which, the content of Ca was the highest (the average value reached 451.2 g/g). Among the determined elements, 15 pairs showed extremely significant positive correlations (P<0.01), and 26 kinds of elements showed significant positive correlations (P<0.05). Through principal component analysis, Al, As, Ba, Ca, Cd, Fe, Mn, Ni, P, Pd, Pb, Se, V, Zn were determined as the characteristic inorganic element ofC.tangutorumO. E. Schulz.

In many plants with anti-cancer effects, the contents of Fe, Mn and Zn were usually higher. Sr, Cr,etc. were also very important[19]. The results of this experiment show that the contents of beneficial trace elements inC.tangutorumO. E. Schulz were higher. Six samples ofC.tangutorumO. E. Schulz were rich in Fe, Mn, Zn and Sr, with a certain amount of Cr. It is speculated that inorganic elements may play an important role in the anticancer effect ofC.tangutorumO. E. Schulz.

This article was established a method for simultaneous determination of 29 kinds of inorganic elements inC.tangutorumO. E. Schulz by set digestion and inductively coupled plasma optical emission spectroscopy (ICP-OES). The contents of inorganic elements in samples ofC.tangutorumO. E. Schulz from six different origins were determined, and the determination results were analyzed, so as to provide a reference basis for the quality control, safety evaluation ofC.tangutorumO. E. Schulz and the development and utilization of big health products of medicinal and edible plants.