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In This Article

  • Summary
  • Abstract
  • Introduction
  • Protocol
  • Representative Results
  • Discussion
  • Acknowledgements
  • Materials
  • References
  • Reprints and Permissions

Summary

Here, a headspace gas chromatography-tandem quadrupole mass spectrometry (HS-GC-MS/MS) method suitable for the determination of trimethylamine (TMA) in animal-derived medicines is described. The protocol includes sample pretreatment, headspace treatment, analysis conditions, methodological validation, and the determination of TMA in animal-derived medicines.

Abstract

Animal-derived medicines have distinctive characteristics and significant curative effects, but most of them have an obvious fishy odor, resulting in the poor compliance of clinical patients. Trimethylamine (TMA) is one of the key fishy odor components in animal-derived medicine. It is difficult to identify TMA accurately using the existing detection method due to the increased pressure in the headspace vial caused by the rapid acid-base reaction after the addition of lye, which causes TMA to escape from the headspace vial, stalling the research progress of the fishy odor of animal-derived medicine. In this study, we proposed a controlled detection method that introduced a paraffin layer as an isolation layer between acid and lye. The rate of TMA production could be effectively controlled by slowly liquefying the paraffin layer through thermostatic furnace heating. This method showed satisfactory linearity, precision experiments, and recoveries with good reproducibility and high sensitivity. It provided technical support for the deodorization of animal-derived medicine.

Introduction

Treating human diseases by utilizing products derived from animal parts and/or their by-products (referred to here as animal-derived medicines) is receiving increased attention. They play an important role in treating cancer, cardiovascular disease, liver cirrhosis, mastitis, and other diseases, with the advantages of a strong effect, small dosage, and significant and specific clinical efficacy. However, animal-derived medicines generally have a prominent fishy odor, which greatly affects patients' compliance, and are especially unfavorable for children1,2. The fishy odor mainly comes from the proteins, am....

Protocol

See Table 1 for information on the medicinal materials of Pheretima, Periplaneta americana, and Hirudo. They were identified by Prof. Xu Runchun, Chengdu University of Traditional Chinese Medicine, as the dried bodies of Pheretima aspergillum (E.Perrier), Periplaneta americana L., and Whitmania pigra Whitman.

1. Specimen extraction

  1. Crush Pheretima, Periplaneta americana, and

Representative Results

Schematic diagrams of the pre-processing principle and operation of this protocol are shown in Figure 1 and Figure 2, respectively. The peak time of TMA was 2.3 min, with a sharp peak shape and no interference from other impurities (Figure 3). Measuring the linear range of 0.1-10 µg/mL TMA standard solution, with TMA concentration as the abscissa and peak area as the ordinate, a standard curve was drawn. The linear regression e.......

Discussion

Animal-derived medicines come from the whole body, organs or tissues, physiological or pathological products, excretions or secretions, and processed products of animals. TMA is an important source of fishy odor in animal-derived medicines; it is a typical malodorous substance with a very low olfactory threshold (0.000032 × 10-6 V/V) and a strong fishy odor13. At present, the commonly used HS-GC-MS method cannot detect TMA in animal-derived medicines stably and accurately.

Acknowledgements

This work was supported by grants from the National Natural Science Foundation of China (82173991), and Sichuan Science and Technology Program (2022YFS0442).

....

Materials

NameCompanyCatalog NumberComments
CentrifugeBeckman Coulter Trading (China) Co.SSC-2-0213
Chinese herbal medicine grinderZhejiang Yongkang Xi'an Hardware and Pharmaceutical FactoryHX-200K
Convection ovenSanyo Electric Co., LtdMOV-112F
Decapper for 20 mm Aluminum capsANPEL Laboratory Technologies (Shanghai) IncV1750004
Electronic balanceShimadzu Corporation JapanAUW220D
Gas chromatography mass spectrometryShimadzu Corporation JapanTQ-8050 NX
Headspace VialANPEL Laboratory Technologies (Shanghai) Inc25760200
HomogenizerShanghai biaomo FactoryFJ200-SH
Preassembled CapANPEL Laboratory Technologies (Shanghai) IncL4150050
Sample sieveZhenxing Sieve Factory/
SH-Volatile AmineChengdu Meimelte Technology Co., Ltd227-3626-01
Sodium hydroxideChengdu Chron Chemicals Co., Ltd2022101401
Solid paraffin waxShanghai Hualing Kangfu apparatus factory20221112
Trichloroacetic acidChengdu Chron Chemicals Co., Ltd2022102001
Trimethylamine hydrochlorideChengdu Aifa Biotechnology Co., LtdAF22022108
Ultra-pure water systemSichuan Youpu Ultrapure Technology Co., LtdUPR-11-5T

References

  1. Fan, H., et al. Material basis of stench of animal medicine: a review. China Journal of Chinese Materia Medica. 47 (20), 5452-5459 (2022).
  2. Deng, Y. J., et al. Progress on formation and taste-masking technology of stenc....

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TrimethylamineHeadspace Gas ChromatographyTandem Quadrupole Mass SpectrometryAnimal derived MedicineFishy OdorPretreatment MethodDetection TechniqueParaffin LayerLinearityPrecisionRecoveryReproducibilitySensitivity

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