Arsenic (As) is a ubiquitous element in nature. Common forms of arsenic include arsenite (As(Ⅲ)), arsenate (As(Ⅴ)), monomethylarsenic (MMA), dimethyl arsenic arsenic acid (DMA), arsenic betaine (AsB), arsenic choline (AsC). The toxicity of arsenic depends on the chemical form in which it exists. The more toxic form is inorganic arsenic (As(III)>As(V)). Among organic arsenic, MMA is more toxic than DMA. AsB and AsC are generally considered non-toxic. Different forms of As can cause multi-organ cytotoxicity to varying degrees , causing irreversible damage to the gastrointestinal tract, liver, cardiovascular system, etc., and are also closely related to the formation of various cancers such as skin cancer, lung cancer, liver cancer, etc. Different forms of arsenic have different effects on the pharmacology of Chinese herbal medicines and on the human body. Therefore, it is very necessary to analyze the different forms of arsenic in Chinese herbal medicines.

This experiment was partially improved on the basis of "Chinese Pharmacopoeia 2322 Method for determination of elemental forms and valence states of mercury and arsenic". The method of liquid chromatography-inductively coupled plasma mass spectrometry (LC-ICP-MS) was used to determine the medicinal stone. The content of six different forms of arsenic compounds in calamus was tested. This method has the characteristics of strong applicability, high sensitivity and remarkable separation effect, and can effectively and quickly separate and determine trace amounts of arsenic compounds in different forms.

Experimental part

instrument

Table 1 Liquid Chromatography-Inductively Coupled Plasma Mass Spectrometer

Table 2 Detection parameters of inductively coupled plasma mass spectrometer

Reagents and standards

First-grade water: 18.2 MΩ·cm deionized water;

Ammonia: 25% ~ 28%, electronic grade;

Nitric acid: 67~70%, G3 level;

Ammonium dihydrogen phosphate: ≥99.0%, chromatography grade;

Pepsin (pig gastric mucosa): protease activity ≥250 units/mg, chromatography grade;

Trivalent arsenic, pentavalent arsenic, monomethyl arsenic, dimethyl arsenic, arsenic betaine, arsenic choline solution standard materials.

Sample preparation

After crushing the Acorus medicinal materials, sieve to obtain a 200-mesh Acorus powder sample. Weigh the Acorus calamus powder sample, add the extract, place it in a water bath at 40 degrees Celsius, heat and oscillate for 20 hours, take it out, shake it well, and let it stand for 2 hours. Take the supernatant, filter it with a disposable filter membrane, and put it on the machine for testing. Prepare three sets of parallel sample solutions, named sample-01, sample-02, and sample-03. Prepare a blank solution in the same way.

Results and discussion

chromatographic separation

Figure 1 shows the chromatographic separation spectrum of the 20 μg/L mark. It can be seen from the figure that the six forms have reached baseline separation. The retention times of each form are shown in Table 4.

Figure 1 Chromatogram of arsenic species standard sample

Table 4 Retention times of six arsenic species

Precision test

Three sample solutions were measured. The sample data and precision are shown in Table 6. It can be seen from the table that RSD ≤ 1.2%.

Table 6 Sample precision test results

Spiked recovery test

The samples were tested for spike recovery rate. Each sample was spiked at an appropriate concentration according to the element form content. The spiked samples were prepared in triplicate. The results are shown in Table 7. The spike recovery rate of each sample was between 106% and 106%. Between 110%, RSD≤3.0%

Table 7 Sample spike recovery test results

Extraction rate

Using the above method, the total arsenic and six arsenic forms in the Acorus sample were measured. The total arsenic content in the Acorus sample was 1683 μg/kg and 1593 μg/kg respectively. The calculated extraction rate was 94.7%, indicating that the six species measured above The method has good extraction yields for arsenic species.

in conclusion

This experiment uses a liquid chromatography-inductively coupled plasma mass spectrometry method to test the content of six different forms of arsenic compounds in the traditional Chinese medicine Shichangpu. This method can test the six arsenic forms in the traditional Chinese medicine Shichangpu within 20 minutes. After separation, the linear correlation coefficient values of the six arsenic forms are all greater than 0.999. The above method can effectively separate the six arsenic forms and quickly and accurately determine trace amounts of AsC, AsB, As(Ⅲ), DMA, MMA, As(Ⅴ).

Equipment and consumables solutions

1. Configuration details of SUPEC 7000 liquid chromatography system

2. Reagents and standards