Research Progress in New Techniques and Methods for Identification of Traditional Chinese Medicine

The identification of traditional Chinese medicine (TCM) serves as the foundation of TCM research. In traditional TCM identification, herbal materials are primarily identified through methods such as shape, color discrimination, and odor. However, with the continuous advancement of science and technology, traditional TCM identification methods have become insufficient to meet the needs of modern TCM identification. As a result, TCM identification methods have been continuously updated, giving rise to many new techniques and methods, including DNA molecular marker methods, biosensors, chromatography-mass spectrometry (LC-MS), spectroscopic techniques, and more. These new technologies not only provide scientific evidence for TCM identification, but also promote the development of TCM identification methods.

Related Reading: "Traditional Identification Methods of Chinese Medicines and Their Development and Application"

1. DNA Molecular Identification Technology

DNA molecular identification technology is one of the most common molecular identification techniques. In the process of traditional Chinese medicine identification, it can effectively react to specific DNA fragments in biological individuals. This identification method is highly accurate and is not easily influenced by many external factors, thus accurately reflecting the actual situation of traditional Chinese medicinal materials. DNA molecular identification boasts advantages such as speed, accuracy, and efficiency. Its detection techniques include polymerase chain reaction (PCR), gene chips, and microarray chips. In recent years, with the development of PCR technology and chip technology, DNA molecular identification technology has made significant progress and has been widely applied in the identification of traditional Chinese medicine, especially DNA barcoding technology. Due to its rich DNA sequence information, accuracy, and lack of species restrictions, DNA barcoding technology has been widely used in the identification of traditional Chinese medicinal material varieties. However, the research on DNA barcoding technology is still in its infancy and requires further improvement and development. Currently, commonly used DNA molecular identification techniques include random amplified polymorphic DNA (RAPD) and PCR.

2.Protein Marker Technology

Protein marker technology includes identification techniques such as protein electrophoresis, isoenzyme analysis, protein flight time, and antisera. Protein electrophoresis is commonly used for the identification of fruit seeds and animal-derived medicinal materials. It utilizes the differences in charges, shapes, and sizes of protein molecules within medicinal materials to conduct electrophoretic analysis, effectively identifying different types of traditional Chinese medicinal materials. Isoenzyme analysis is based on the fact that different types of traditional Chinese medicinal materials generally contain different isoenzyme molecular structures, and their immunological activities also vary. Therefore, isoenzyme technology can be used to identify the isoenzyme molecular structures in traditional Chinese medicinal materials, enabling the identification of different medicinal materials based on their differences. Protein flight time mass spectrometry is a commonly used method for identifying the quality fingerprint of traditional Chinese medicinal materials containing protein/peptide components derived from animals. Antisera identification technology primarily focuses on identifying antigenic substances such as proteins in medicinal materials. Many traditional Chinese medicinal materials contain antigenic substances such as polysaccharides and proteins. Using antisera identification technology with protein markers, these antigenic substances can be utilized to prepare corresponding specific substances. Then, through enzyme-linked immunosorbent assay (ELISA), the substances can be tested, accurately reflecting the actual conditions and facilitating clinical identification and application.

3. Biosensors

A biosensor is a detection instrument that utilizes the interaction between biomolecules to detect the content of a substance to be tested. Compared to traditional detection methods, biosensors are characterized by their high sensitivity, ease of operation, non-invasive nature, and low cost. Therefore, biosensors have also gained widespread application in the identification of traditional Chinese medicinal materials. The main technologies based on biosensors include immunological detection technology and molecular hybridization technology. Immunological detection technology primarily involves binding the substance to be tested with antigens or antibodies to form a corresponding complex, and then using immobilized antigens or antibodies to quantitatively detect it. Molecular hybridization technology, on the other hand, utilizes the specific binding of the substance to be tested with related molecules to form a complex, leveraging intermolecular interactions to determine the content of the substance to be tested.

4. Chromatography Technology

High-performance liquid chromatography (HPLC) technology demonstrates a high degree of separation effectiveness in the identification of traditional Chinese medicinal materials. It can effectively distinguish the quantity and characteristics of small-molecule chemical components in medicinal materials, as well as eliminate interference factors through the use of different solvents and extraction methods. Trace quantities can be enriched and presented, enabling the acquisition of multiple molecular chemical fingerprint characteristics of traditional Chinese medicinal materials. However, HPLC requires a large number of medicinal material samples, and different fixed phases, mobile phases, and detection methods are required for different medicinal materials, making it less versatile in identification methods. Additionally, instrument-related factors can also influence the results. HPLC is highly effective in the identification of traditional Chinese medicine decoctions.

Gas chromatography is based on the analysis of medicinal materials based on the different distribution of compounds between a fixed gas flow and a fixed phase. It is commonly used clinically for detecting gaseous, volatile, and semi-volatile substances. Programmed temperature rise is the primary method for identifying gas chromatography. Gas chromatography primarily identifies thermally stable and volatile chemical components in traditional Chinese medicine, serving as a complement to HPLC. However, this identification method lacks standardized chromatographic conditions, resulting in suboptimal results.

Chromatography-mass spectrometry (chromatography-MS) is a rapid analytical technique that combines liquid chromatography, gas chromatography, and mass spectrometry. The application of chromatography-MS has provided a new method for the identification of traditional Chinese medicine, significantly advancing the development of this field. However, this method also has certain limitations, such as small sample sizes, slow analysis speed, and complex sample preparation, which restrict its application in the identification of traditional Chinese medicine.

5. Spectral Analysis

Spectral technology is an analytical method that involves the absorption of light by a substance at different wavelengths. Through mathematical calculations and processing, the physical quantity of the substance to be measured is converted into spectral information, and its characteristics are described in the form of spectra. It can be used not only to study the structure of substances but also to identify and authenticate their components. Spectral analysis has the advantages of fast detection speed, short analysis cycle, wide applicability, and simple operation. Currently, the main spectral analysis methods include ultraviolet-visible spectrophotometry (UV), fluorescence spectrophotometry (FL), infrared spectroscopy (IR), nuclear magnetic resonance (NMR), molecularly imprinted polymer fingerprint spectrum technology (MAS), and ultra-performance liquid chromatography-diode array detection (UPLC-DA). Spectral technology is widely used in the identification of traditional Chinese medicine, where it can authenticate the authenticity and quality of medicinal materials. For example, infrared spectroscopy can be used to identify traditional Chinese medicinal materials from different origins, and ultraviolet-visible spectrophotometry can be used to distinguish between genuine and inferior medicinal materials. 

6. Fingerprint Spectrum

Fingerprint spectrum refers to the common chemical components and identical chromatographic peaks existing between different medicinal materials or different parts of the same medicinal material. By utilizing these common chemical components and chromatographic peaks, and through a certain mathematical model calculation, one can distinguish these common chemical components from other components contained in the medicinal material, thus facilitating the identification of traditional Chinese medicine. Fingerprint spectrum is one of the important methods for the identification and quality control of traditional Chinese medicinal materials, mainly including chromatographic fingerprint spectrum, microscopic fingerprint spectrum, and bioactive fingerprint spectrum. Among them, chromatographic fingerprint spectrum is a primary method for the identification of traditional Chinese medicinal materials and is also the most widely used method in traditional Chinese medicine identification. Chromatographic fingerprint spectrum technology has the advantages of simple operation, rapidity, good reproducibility, and strong versatility, and it has been widely applied in the identification of traditional Chinese medicine. With the continuous advancement of chromatography technology, its application in traditional Chinese medicine identification has become increasingly widespread, such as chemical composition fingerprint spectrum of traditional Chinese medicine, chromatography-mass spectrometry technology, molecular imprinting technology, and mass spectrometry technology. Through the combined application of different methods, traditional Chinese medicinal materials of different types and origins can be distinguished and identified.

The traditional methods for identifying traditional Chinese medicinal materials mainly rely on observing the shape of the herbs visually, feeling them by hand, and smelling their odor to determine their authenticity and quality. However, these methods are easily influenced by subjective factors, and the accuracy cannot be guaranteed. New methods of identifying traditional Chinese medicine based on the molecular level have achieved some results in authenticating traditional Chinese medicinal materials. DNA barcoding technology has been widely applied in species identification of traditional Chinese medicinal materials. By establishing a DNA barcode database of standard sequences for traditional Chinese medicinal materials, as well as screening, obtaining, and detecting marker genes, the standardization of species identification of traditional Chinese medicinal materials can be gradually achieved. At the same time, with the establishment of fingerprint spectrum technology and quality control standard system for traditional Chinese medicine, the quality and safety of traditional Chinese medicinal materials have also been further ensured.

References

[1] Luo Lun, Ran Huimin. A Brief Discussion on the Research Progress of New Technologies and Methods in the Identification of Traditional Chinese Medicine [J]. Journal of Clinical Medical Literature Electronic Edition, 2019, 6(37): 189+191.

[2] Zhou Zhengli. Research Progress on New Technologies and Methods of Chinese Medicine Identification [J]. Chinese Science and Technology Journal Database (Medical Sciences), 2023, Issue 

Author's Introduction

Xiaomichong, a researcher in drug quality, has long been committed to drug quality research and validation of drug analysis methods. Currently, he/she is employed at a large domestic pharmaceutical research and development company, engaged in drug inspection analysis and validation of analytical methods.