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Construction method and application of earthworm 1H NMR fingerprint spectrum

A fingerprint and construction method technology, applied in the field of construction of Dilong 1HNMR fingerprint, can solve the problems of inability to clarify the quality difference of Dilong, difficult to retain and detect, and unable to objectively and accurately compare the quality uniformity of Dilong decoction pieces

Inactive Publication Date: 2020-06-12
SHANXI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Small molecular substances such as free amino acids in Earthworm are closely related to their anticoagulant activity. Most of these components have no UV absorption and are highly polar water-soluble substances, which are difficult to retain and detect on conventional C18 chromatographic columns
In addition, only the chromatographic peak corresponding to hypoxanthine is known in the fingerprint, and the rest of the chromatographic peaks are unknown, so it is impossible to clarify which components cause the quality difference of earthworm
In summary, none of the existing methods can objectively and accurately compare the quality uniformity of different batches of Dilong decoction pieces

Method used

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  • Construction method and application of earthworm 1H NMR fingerprint spectrum
  • Construction method and application of earthworm 1H NMR fingerprint spectrum
  • Construction method and application of earthworm 1H NMR fingerprint spectrum

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] A method for constructing earthworm 1H NMR fingerprints in this embodiment comprises the following steps:

[0028] Step 1, take 100 mg of earthworm powder and add it to 6 mL of 20% methanol-water mixed solution, vortex for 1 minute, ultrasonically extract for 10 minutes, centrifuge, remove supernatant and spin dry to obtain earthworm extract;

[0029] Step 2, dissolve the earthworm extract with 600 μL of a mixed solution of deuterated methanol and heavy water (the volume ratio of deuterated methanol and heavy water is 1:1), transfer to a centrifuge tube, 13000r min -1 Centrifuge for 20 minutes, pipette 600 μL of the supernatant into an NMR tube for NMR testing, import the spectrum into the NMR data processing software, and obtain the 1H NMR fingerprint of the earthworm medicinal material;

[0030] Step 3, identify the 1H NMR spectrum of Earthworm, and determine the characteristic peaks of the fingerprint spectrum of Earthworm by using the reference substance combined wi...

Embodiment 2

[0032] A method for constructing earthworm 1H NMR fingerprints in this embodiment comprises the following steps:

[0033] Step 1, take 300mg of earthworm powder and add it to 6mL of 60% methanol-water mixed solution, vortex for 1min, ultrasonically extract for 30min, centrifuge, remove supernatant and spin dry to obtain earthworm extract;

[0034] Step 2, dissolve the earthworm extract with 1000 μL of a mixed solution of deuterated methanol and heavy water (the volume ratio of deuterated methanol and heavy water is 1:1), transfer to a centrifuge tube, 13000r min -1 Centrifuge for 20 minutes, pipette 600 μL of the supernatant into an NMR tube for NMR testing, and import the spectrum into the NMR data processing software to obtain the 1H NMR fingerprint of the earthworm medicinal material;

[0035] Step 3, identify the 1H NMR spectrum of Earthworm, and determine the characteristic peaks of the fingerprint spectrum of Earthworm by using the reference substance combined with the c...

Embodiment 3

[0037] A method for constructing earthworm 1H NMR fingerprints in this embodiment comprises the following steps:

[0038] Step 1, take 500mg of earthworm powder and add it to 6mL of 50% methanol-water mixed solution, vortex for 1min, ultrasonically extract for 25min, centrifuge, remove supernatant and spin dry to obtain earthworm extract;

[0039] Step 2, dissolve the earthworm extract with 800 μL of a mixed solution of deuterated methanol and heavy water (the volume ratio of deuterated methanol and heavy water is 1:1), transfer to a centrifuge tube, 13000r min -1 Centrifuge for 20 minutes, pipette 600 μL of the supernatant into an NMR tube;

[0040] Spectrum collection: The sample was measured on a 600MHz NMR instrument at 25°C, the measurement frequency was 600.13MHz, the pulse sequence was noesygppr1d, the number of scans was 64, the spectrum width was 12345.7Hz, and the sampling data points were 65536.

[0041] Spectrum preprocessing: NMR spectra were imported into MestReNo...

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Abstract

The invention belongs to the technical field of fingerprint spectrum construction, and particularly relates to a construction method and application of an earthworm 1H NMR fingerprint spectrum. According to the invention, an earthworm fingerprint spectrum and the characteristic peak are obtained by adopting a nuclear magnetic resonance technology; 29 chemical components in the earthworm can be represented at the same time; the invention has the advantages that various metabolites including amino acids, organic acids, nucleotides and the like in the earthworm decoction pieces are not limited byultraviolet absorption of substances and retention of chromatographic columns, the chemical components and quality of earthworms can be comprehensively reflected, and the method can be used for controlling the quality uniformity of the earthworm decoction pieces through multivariate statistical analysis and univariate analysis of the identified substances.

Description

technical field [0001] The invention belongs to the technical field of fingerprint atlas construction, and in particular relates to a construction method and application of earthworm 1H NMR fingerprint atlas. Background technique [0002] Earth Dragon is derived from the dried bodies of Pheretima aspergillum (E.Perrier), Pheretima vu1garis Chen, Pheretima guillelmi (Michaelsen) or Pectinifera Michaelsen of the family Pheretima aspergillum. The former is commonly called "Guangdilong", and the latter three are commonly called "Shanghaidilong". Guangdilong is caught from spring to autumn, Hudilong is caught in summer, the abdomen is cut open in time, viscera and sediment are removed, washed, and dried in the sun or at low temperature. [0003] Dilong is a traditional insect medicine in my country. It has the effects of clearing away heat, calming convulsions, dredging collaterals, relieving asthma, and diuresis. It has clinically obvious anticoagulant activity. Earthworm cont...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/86G01N30/64G01N30/06
CPCG01N30/02G01N30/06G01N30/64G01N30/8686G01N2030/065
Inventor 何盼李震宇秦雪梅
Owner SHANXI UNIV