CN115246873A - Syngnathus characteristic polypeptide, application thereof and method for identifying Syngnathus - Google Patents

Syngnathus characteristic polypeptide, application thereof and method for identifying Syngnathus Download PDF

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CN115246873A
CN115246873A CN202210653950.9A CN202210653950A CN115246873A CN 115246873 A CN115246873 A CN 115246873A CN 202210653950 A CN202210653950 A CN 202210653950A CN 115246873 A CN115246873 A CN 115246873A
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syngnathus
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尹雪
焦阳
林永强
汪冰
于雅萌
解盈盈
周倩倩
梅桂雪
牛艳
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Shandong Institute for Food and Drug Control
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Abstract

The invention relates to a syngnathus pseudopterus characteristic polypeptide, application thereof and a method for identifying the syngnathus pseudopterus, belonging to the technical field of biotechnology detection. The invention provides two unique characteristic polypeptides of syngnathus, and the amino acid sequences of the characteristic polypeptides are shown in SEQ ID NO.1 and SEQ ID NO. 2. The characteristic polypeptide provided by the invention has excellent specificity and stability for the syngnathus mimicus, has strong specificity, can be used for identifying the syngnathus mimicus, and has good application prospect.

Description

Syngnathus characteristic polypeptide, application thereof and method for identifying Syngnathus
Technical Field
The invention relates to the technical field of biotechnology detection, in particular to two Syngnathus characteristic polypeptides, application thereof and a method for identifying Syngnathus.
Background
The Syngnathus is an important animal medicinal material in China, has the effects of warming kidney, tonifying yang, resolving masses and relieving swelling, and has long administration history and numerous primordial sources. The pharmaceutical base source of Syngnathus is defined in the Chinese pharmacopoeia as dried bodies of Syngnathus caudatum hardwickii (Gray), syngnathus biaceutus (Bloch) or Syngnathus acus cusa cusLinnaeus, which are Syngnathus animals of the Syngnaceae family.
The sea dragon as a tonifying medicinal material has an increasing market demand year by year, and the price of the sea dragon in the market is gradually increased because the sea dragon cannot effectively realize artificial breeding and the wild resource of the sea dragon is gradually reduced at present. The sea dragon is not only used as a raw material and directly crushed or extracted to prepare a Chinese patent medicine, but also is often prepared into sea dragon tonic wine to be used as a health-care tonic product.
The species identification and quality control methods of the current Syngnathus mainly comprise a sex identification method, an HPLC fingerprint spectrum method and a molecular biology method. The character identification method generally requires that the sea dragon individual is complete, and the person needing to be identified has rich experience, and the method has certain subjectivity; the specificity of an HPLC fingerprint spectrum method is low; the molecular biology method has high specificity, but is limited by the intact preservation degree of genetic materials in samples, and for the samples of the Syngnathus medicated liquor, the processed products, the extracts, the worm-eaten mildew and a plurality of individual mixed samples, the successful experiment can not be carried out due to the difficulty in extracting DNA. In recent years, more and more methods and standards have been used to identify or quantify natural drugs using characteristic peptide fragments. The protein contained in each species has difference of amino acid at individual site, and the difference site can be intercepted by using proper protease to be used as polypeptide segment with identification meaning, i.e. characteristic peptide segment. The detection of the characteristic peptide fragment generally uses a mass spectrum means, protein activity does not need to be considered, the preparation method is simple, and the property of the peptide fragment is relatively stable. The existing method for identifying the Syngnathus has more limiting factors, can not be applied to derivative products such as Syngnathus powder, syngnathus medicated liquor, processed products, extracts and the like, and the characteristic peptide segment in the Syngnathus protein has potential to be used as an index component for identifying the species of the Syngnathus. At present, no Syngnathus-related protein sequence information is collected in each database, and a differential peptide segment cannot be obtained through protein sequence comparison, so that no related records of Syngnathus-characteristic polypeptides exist.
Disclosure of Invention
The invention aims to provide two quasi-sea dragon characteristic polypeptides, application thereof and a method for identifying the quasi-sea dragon. The characteristic polypeptide provided by the invention has excellent specificity and stability for the syngnathus, has strong specificity, can be used for identifying the species of the medicinal material of the syngnathus, particularly the species of the syngnathus, and has good application prospect.
The invention provides two quasi-sea dragon characteristic polypeptides, wherein the amino acid sequences of the characteristic polypeptides are shown as SEQ ID NO.1 and SEQ ID NO. 2.
The invention also provides application of the characteristic polypeptide in the technical scheme in identifying the Syngnathus.
Further, the application of SEQ ID NO.1 and SEQ ID NO.2 in identifying the Syngnathus independently or together.
Preferably, the identifying is performed using mass spectrometry, and detecting the ion pair comprises: SEQ ID No.1 is a quantitative ion with mass-to-charge ratio of m/z 639.6 → 810.2 and a qualitative ion with mass-to-charge ratio of m/z 639.6 → 753.1; the mass-to-charge ratio of the quantitative ion of SEQ ID NO.2 is m/z852.1 → 1144.6, and the mass-to-charge ratio of the qualitative ion of m/z852.1 → 875.4.
The invention also provides an identification method of the Syngnathus pseudopterus, which adopts the characteristic polypeptide shown in SEQ ID NO.1 and/or SEQ ID NO.2 as a reference substance.
The invention also provides a method for identifying the Syngnathus based on the characteristic polypeptide, which comprises the following steps:
(1) Preparation of a test solution: mixing the pretreated pipefish powder extract or the pretreated pipefish medicinal liquor extract with trypsin respectively to obtain a mixed solution, mixing the mixed solution with an ammonium bicarbonate aqueous solution, carrying out enzymolysis and filtering, and taking a filtrate to obtain a test solution;
(2) Preparation of control solutions: the sea-like dragon characteristic peptide is used as a reference substance, and water is added for dissolving to obtain a reference substance solution;
(3) Detection and analysis: detecting and analyzing by using a triple quadrupole mass spectrometry;
the step (1) or the step (2) is not limited in chronological order.
Preferably, in the step (1), the preparation method of the pretreated pipefish powder extract comprises the following steps: mixing Syngnathus powder with water, decocting for three times (4 h,3h and 2 h), mixing decoctions, and diluting with ammonium bicarbonate water solution to obtain pretreated Syngnathus powder extractive solution;
the preparation process of the pretreated Syngnathus medicinal liquor extract comprises the following steps: taking out Syngnathus in Syngnathus medicated liquor, drying at low temperature, decocting with water for three times (4 hr, 3 hr and 2 hr respectively), mixing decoctions, filtering, and collecting filtrate to obtain pretreated Syngnathus medicated liquor extract;
preferably, the trypsin is added in the form of an aqueous trypsin solution, and the mass concentration of the trypsin in the aqueous trypsin solution is 1mg/ml.
Preferably, the mass percentage content of ammonium bicarbonate in the ammonium bicarbonate aqueous solution is 1%.
Preferably, the enzymolysis condition is enzymolysis at 37 ℃ for 2h.
Preferably, the triple quadrupole mass spectrometry is performed by taking 0.1% formic acid acetonitrile as a mobile phase A and 0.1% formic acid solution as a mobile phase B, and performing gradient elution: 0-3min, 5%; the flow rate was 0.5ml per minute.
Preferably, the triple quadrupole mass spectrometry adopts a mass spectrometry detector and an electrospray positive ion mode to carry out multi-reaction monitoring, wherein the SEQ ID NO.1 selects a quantitative ion with a mass-to-charge ratio of m/z 639.6 → 810.2, the qualitative ion with a mass-to-charge ratio of m/z 639.6 → 753.1 is taken as a detection ion pair, the SEQ ID NO.2 selects a quantitative ion with a mass-to-charge ratio of m/z852.1 → 1144.6, and the qualitative ion with a mass-to-charge ratio of m/z852.1 → 875.4 is taken as a detection ion pair.
The invention provides two polypeptides with characteristics of syngnathus. The characteristic polypeptide provided by the invention is used as an index component for identifying the Syngnathus, can be distinguished from the Syngnathus of other species, and lays a foundation for developing methods for identifying the Syngnathus and related products. Specifically, the characteristic polypeptide of the invention has the following beneficial effects:
(1) The method intercepts the difference polypeptide segments in the protein sequence, adopts mass spectrum for detection, has simple and rapid operation and stable determination result, is not interfered by the shape and the processing process of a Syngnathus sample, and fills the blank of the method for identifying the species of the Syngnathus pseudosea and detecting the Syngnathus in the Syngnathus product;
(2) The characteristic polypeptide and the detection method provided by the invention provide reference for searching characteristic peptide segments in related species lacking in a database;
(3) The characteristic polypeptide provided by the invention has excellent specificity and stability for the syngnathus, has strong specificity, can be used for identifying the medicinal material of the syngnathus, and has good application prospect.
Drawings
FIGS. 1-9 are the specificity maps of the Syngnathus pseudopeptide SEQ ID NO.1 provided by the present invention; wherein FIG. 1 is the M/z 639.6 → 810.2, 753.1 map of SEQ ID NO.1 reference substance, and FIGS. 2-9 are the m/z 639.6 → 810.2, 753.1 maps extracted from Syngnathus pseudosea, cynanchum paniculatum, syngnathus schlegeli, dowanese Syngnathus polycephalus, baojia Syngnathus, elaphe robusta, rhynchosia lucida, phlomis gesii, and Rhynchophorus glehnsonii in sequence.
FIGS. 10-18 are the Syngnathus pseudopeptides SEQ ID NO.2 specificity profiles provided by the present invention; wherein FIG. 10 is the M/z852.1 → 1144.6 and 875.4 maps of SEQ ID NO.2 reference substance, and FIGS. 11-18 are the m/z852.1 → 1144.6 and 875.4 maps of Syngnathus niponicas, cynanchum paniculatum, syngnathus schlegeli, syngnathus polygama, syngnathus galaxacum, syngnathus alurus, syngnathus crassimus, syngnathus platyphylla, and Syngnathus gelloides.
Detailed Description
The invention provides two sea dragon simulating characteristic polypeptides, wherein the amino acid sequence of the characteristic polypeptides is shown as SEQ ID NO.1: NAGPQGPQGAAGPR, SEQ ID NO.2: QTGGAGSPATTGFPSGPGR. The invention provides a peptide fragment with characteristic recognition significance for syngnathus. The method comprises the steps of carrying out trypsin enzymolysis on water extracts of different kinds of Syngnathus such as Syngnathus pseudognathus, cynanchum brachypomum, cynanchum polycephalum, syngnathus schlegeli, etc., and analyzing by using a nanoliter liquid chromatography-high resolution mass spectrometer to obtain a Syngnathus protein complete peptide fragment mass spectrogram; carrying out chemometric analysis on the mass spectrum data, and then carrying out specificity verification on the experimental result by using a triple quadrupole mass spectrum; finally, 2 characteristic ion pairs are provided, corresponding to 2 characteristic peptide fragments, and the specificity verification result is good.
The invention also provides application of the characteristic polypeptide in the technical scheme in identifying the Syngnathus.
In the present invention, the identifying is performed using mass spectrometry, and detecting the ion pair comprises: SEQ ID No.1 is a quantitative ion with mass-to-charge ratio of m/z 639.6 → 810.2 and a qualitative ion with mass-to-charge ratio of m/z 639.6 → 753.1; the mass-to-charge ratio of the quantitative ion with the SEQ ID NO.2 is m/z852.1 → 1144.6, and the mass-to-charge ratio of the qualitative ion with the m/z852.1 → 875.4.
The invention also provides a method for identifying the Syngnathus based on the characteristic polypeptide, which comprises the following steps:
(1) Mixing the pretreated pipefish powder extract or the pretreated pipefish medicinal liquor extract with trypsin respectively to obtain a mixed solution, mixing the mixed solution with an ammonium bicarbonate aqueous solution, carrying out enzymolysis and filtering, and taking a filtrate to obtain a test solution;
(2) Adding an ammonium bicarbonate aqueous solution to dissolve the syngnathus pseudopterus characteristic peptide serving as a reference substance in the technical scheme to obtain a reference substance solution;
(3) Detecting and analyzing by using a triple quadrupole mass spectrometry;
the steps (1) or (2) are not limited in chronological order.
The method provided by the invention intercepts the differential polypeptide segments in the protein sequence, adopts mass spectrum for detection, is simple and rapid to operate, has stable determination result, is not interfered by the form and the processing process of a Syngnathus sample, fills the blank of the method for identifying the Syngnathus pseudopterus species and detecting the Syngnathus in the Syngnathus product, and provides reference for searching the characteristic peptide segments in the kindred species lacking in the database.
Mixing the pretreated pipefish powder extract or the pretreated pipefish medicinal liquor extract with trypsin to obtain a mixed solution, mixing the mixed solution with an ammonium bicarbonate aqueous solution, carrying out enzymolysis and filtering, and taking a filtrate to obtain a test solution. In the present invention, the preparation method of the pretreated pipefish powder extract preferably comprises the following steps: mixing Syngnathus powder with water, decocting for three times (4 h,3h and 2h respectively), and mixing decoctions to obtain pretreated Syngnathus powder extract. The invention preferably selects 1g of Syngnathus powder, and the total amount of the decoction is preferably 50mL. Specifically, the method preferably takes 5mL of the decoction, places the decoction in a 50mL measuring flask, uses an ammonium bicarbonate aqueous solution to dilute the decoction to a scale, and shakes the solution uniformly to obtain the pretreated pipefish powder extracting solution. In the present invention, the preparation process of the pretreated Syngnathus medicated liquor extract preferably comprises the following steps: taking out Syngnathus in Syngnathus medicated liquor, drying at low temperature, decocting with water for three times (4 hr, 3 hr and 2 hr respectively), mixing decoctions, filtering, and collecting filtrate to obtain pretreated Syngnathus medicated liquor extract. After obtaining the pretreated pipefish extract and the pretreated pipefish medicinal liquor extract, the invention preferably measures 1-5 mL pipefish powder extract or pipefish medicinal liquor extract, preferably adds 100 μ L trypsin solution, shakes evenly, adds ammonium bicarbonate aqueous solution to 50mL, shakes evenly, carries out enzymolysis after sealing. In the present invention, the enzymolysis condition is preferably enzymolysis at 37 ℃ for 2h. After enzymolysis, the method is preferably cooled, more preferably to normal temperature, and the temperature range of the normal temperature is preferably 10-30 ℃. After cooling, the invention filters and takes the filtrate as the test solution.
In the invention, the trypsin is added in the form of an aqueous trypsin solution, and the mass concentration of the trypsin in the aqueous trypsin solution is 1mg/ml. In the invention, the mass percentage of ammonium bicarbonate in the ammonium bicarbonate aqueous solution is 1%.
The invention takes the pseudo-sea dragon characteristic peptide as a reference substance, and water is added for dissolution to obtain a reference substance solution. The source of the Syngnathus pseudopeptides is not particularly limited, and the Syngnathus pseudopeptides can be obtained by artificial synthesis, for example, 2 Syngnathus pseudopeptides of the invention are obtained by entrusted Nanjing peptide Biotech limited according to the specified amino acid sequence.
The invention utilizes triple quadrupole mass spectrometry to carry out detection and analysis. In the present invention, the triple quadrupole mass spectrometry is preferably performed by gradient elution with 0.1% formic acid acetonitrile as mobile phase a and 0.1% formic acid solution as mobile phase B: 0 to 3min,5% by weight A → 8% by weight A; the flow rate was 0.5ml per minute. In the invention, the triple quadrupole mass spectrometry is preferably carried out by adopting a mass spectrometry detector in an electrospray positive ion mode, and multiple reactions are monitored, wherein the SEQ ID NO.1 selects a quantitative ion with a mass-to-charge ratio of m/z 639.6 → 810.2 and a qualitative ion with a mass-to-charge ratio of m/z 639.6 → 753.1; SEQ ID NO.2 selects the quantitative ion with mass/charge ratio m/z852.1 → 1144.6, the qualitative ion with m/z852.1 → 875.4.
The two polypeptides with characteristics of Syngnathus, the application thereof and the method for identifying Syngnathus are described in detail in the following examples, but the technical scheme of the invention includes but is not limited to the following examples.
Triple quadrupole mass spectrometry conditions
Gradient elution was carried out using 0.1% formic acid acetonitrile as mobile phase A and 0.1% formic acid solution as mobile phase B (0-3min, 5%. A → 8%. A); the flow rate was 0.5ml per minute. Performing Multiple Reaction Monitoring (MRM) by using a mass spectrum detector and an electrospray positive ion mode (ESI +), wherein the mass charge ratio of quantitative ions with m/z 639.6 → 810.2 and the mass charge ratio of qualitative ions with m/z 639.6 → 753.1 are selected from SEQ ID NO. 1; SEQ ID NO.2 selects the quantitative ion with mass/charge ratio m/z852.1 → 1144.6, the qualitative ion with m/z852.1 → 875.4.
Example 1: quasi-sea dragon characteristic peptide segment search
(1) Preparation of test solution
Sea dragon water extract: taking 100g of a test sample Syngnathus, crushing, putting into a triangular flask, adding water, decocting for three times, namely 4h,3h and 2h, combining the decoctions to be 500mL, slightly boiling, concentrating to be viscous, transferring into a silica gel bowl, and drying in an electric heating constant-temperature air drying oven at 60 ℃ until the Syngnathus water extract is solid to obtain the Syngnathus water extract;
test solution: weighing 0.1g of Syngnathus aqueous extract sample, adding 50ml of 1% ammonium bicarbonate aqueous solution, performing ultrasonic treatment for 30min for dissolving, filtering with microporous membrane, collecting 100 μ L of filtrate, adding 10 μ L of 1mg/ml trypsin aqueous solution, performing enzymolysis at 37 deg.C for 2h, taking out, and cooling to room temperature.
(2) Selection of characteristic ions and sequence estimation
After the test solution is analyzed by nanoliter liquid chromatography-high resolution mass spectrometry, mass spectrum data is imported into PEAKS 8.5 software, de novo sequencing and sequence prediction of all peptide segments of the sample are carried out, and the obtained result is analyzed. And selecting peptide fragments which can be detected only in the Syngnathus sp and are hardly detected in other species of Syngnathus sp as parent ions from the analysis results, analyzing the secondary spectra one by one, and selecting daughter ions with better response. Further analysis yielded mass spectral ion pair information representing the variability of Syngnathus m/z 639.6 double charge) → 810.2, 753.1 and m/z852.1 (double charge) → 1144.6, 875.4. Through further analysis, the characteristic peptide fragment sequence of the syngnathus sp corresponding to the ion is presumed, and the sequence shown in SEQ ID NO.1: NAGPQGPQGAAGPR, SEQ ID NO.2: QTGGAGSPATTGFPSGPGR.
And (3) entrusting and synthesizing a quasi-sea dragon characteristic peptide reference substance according to the presumed amino acid sequence, simultaneously detecting the reference substance and the quasi-sea dragon test sample solution, and confirming the correctness of the sequence by keeping the retention time of the reference substance and the quasi-sea dragon test sample solution consistent with the second-level mass spectrum information.
Example 2: research on specificity of Syngnathus pseudopeptides
The water extracts of all species of the syngnathus were detected by triple quadrupole mass spectrometry, and the results showed that all three species of the syngnathus exhibited a characteristic peptide chromatographic peak, while none of the other multi-swallow syngnathus, shushi syngnathus, baojia gungsonian, jonghui syngnathus, and gelsemium were associated with a chromatographic peak. See fig. 1-9 and fig. 10-18. The characteristic peptide is specific to the Syngnathus, and can be used as an index component for identifying the Syngnathus.
Example 3: application of syngnathus characteristic peptide in syngnathus powder extract and syngnathus medicinal liquor
(1) Sample preparation
Preparing a test solution of a syngnathus powder extracting solution: taking 1g of a test sample, crushing, putting into a triangular flask, adding water, decocting for three times, respectively 4h,3h and 2h, combining decoction liquids, weighing 50mL in total, weighing 1mL to 5mL of measuring flask, adding 100 mu l of 1mg/mL trypsin aqueous solution, shaking up, adding 1% ammonium bicarbonate aqueous solution to a constant volume to scale, shaking up, sealing, carrying out enzymolysis at 37 ℃ for 2h at constant temperature, cooling, filtering, and taking a subsequent filtrate as a test sample solution.
Preparing a test solution of the Syngnathus medicinal liquor: taking 10g of Syngnathus in the Syngnathus medicated liquor, drying at low temperature, decocting with water for three times, respectively for 4h,3h and 2h, and mixing the decoctions to obtain 50mL; filtration was carried out, and the filtrate was taken in 1ml to 5ml measuring bottles and prepared as above from "100. Mu.l of 1mg/ml trypsin aqueous solution was added".
Quasi-sea dragon characteristic peptide fragment reference substance solution: respectively weighing 10mg of each of the quasi-Syngnathus characteristic peptide reference substances SEQ ID NO.1 and SEQ ID NO.2, placing in a 50ml measuring flask, adding water to dissolve, and metering to a certain volume to obtain the final product.
(2) The measurement was performed under triple quadrupole mass spectrometry conditions.
(3) Sample assay
And (3) measuring 10 collected quasi-sea dragon powder extract samples and 5 collected quasi-sea dragon medicinal liquor samples, and detecting quasi-sea dragon characteristic peptides in results, wherein the two characteristic peptides can be used for identifying quasi-sea dragon components in sea dragon products.
Example 4: ion information with discrimination potential
5 pieces of ion information with identification potential are screened from 350 candidate ions, triple quadrupole mass spectrometry verifies that the identification of the pseudo-pipefish component can not be realized by any ions, the specificity of part of ions to the pseudo-pipefish is insufficient, the specificity of ions with the mass-to-nuclear ratio of 639.6 and 852.1 is good, and specific results are shown in table 1.
TABLE 1 results of the specificity verification
Figure BDA0003688465730000081
The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and decorations can be made without departing from the principle of the present invention, and these modifications and decorations should also be regarded as the protection scope of the present invention.
Sequence listing
<110> Shandong province food and drug inspection research institute
<120> quasi-sea dragon characteristic polypeptide, application thereof and method for identifying quasi-sea dragon
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Asn Ala Gly Pro Gln Gly Pro Gln Gly Ala Ala Gly Pro Arg
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Gln Thr Gly Gly Ala Gly Ser Pro Ala Thr Thr Gly Phe Pro Ser Gly
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Pro Gly Arg

Claims (10)

1. A sea dragon-like characteristic polypeptide is characterized in that the amino acid sequence of the characteristic polypeptide is shown as SEQ ID NO.1 and/or SEQ ID NO. 2.
2. Use of the polypeptide according to the characteristics of claim 1 for identifying a Syngnathus.
3. Use according to claim 2, wherein said identification is carried out using mass spectrometry, and wherein the detection of ion pairs by SEQ ID No.1 comprises: quantitative ions with mass-to-charge ratio of m/z 639.6 → 810.2, and qualitative ions with mass-to-charge ratio of m/z 639.6 → 753.1; the detection ion pair of SEQ ID NO.2 comprises: quantitative ions with mass-to-charge ratio of m/z852.1 → 1144.6, and qualitative ions with mass-to-charge ratio of m/z852.1 → 875.4.
4. A method for identifying Syngnathus sp, which comprises using the polypeptide of claim 1 as a reference.
5. The authentication method according to claim 4, comprising the steps of:
(1) Preparing a test solution: mixing the pretreated pipefish powder extract or the pretreated pipefish medicinal liquor extract with trypsin respectively to obtain a mixed solution, mixing the mixed solution with an ammonium bicarbonate aqueous solution, carrying out enzymolysis and filtering, and taking a filtrate to obtain a test solution;
(2) Preparation of control solutions: respectively dissolving the Syngnathus pseudopterus characteristic peptide of claim 1 in water to obtain reference solutions;
(3) Detection and analysis: detecting and analyzing by using a triple quadrupole mass spectrometry;
the steps (1) or (2) are not limited in chronological order.
6. The method according to claim 5, wherein in step (1), the preparation method of the pretreated Syngnathus powder comprises the steps of: mixing Syngnathus powder with water, decocting for three times (4 h,3h and 2 h), mixing decoctions, and diluting with ammonium bicarbonate water solution to obtain pretreated Syngnathus powder extractive solution;
the preparation process of the pretreated pipefish medicinal liquor extracting solution comprises the following steps: taking out Syngnathus in Syngnathus medicated liquor, drying at low temperature, decocting with water for three times (4 h,3h and 2h respectively), mixing decoctions, filtering, and collecting filtrate to obtain pretreated Syngnathus medicated liquor extract.
7. The method according to claim 5, wherein the trypsin is added in the form of an aqueous trypsin solution, wherein the mass concentration of the trypsin in the aqueous trypsin solution is 1mg/ml.
8. The method of claim 5, wherein the aqueous ammonium bicarbonate solution comprises 1% ammonium bicarbonate by weight; the enzymolysis condition is enzymolysis for 2 hours at 37 ℃.
9. The method of claim 5, wherein the triple quadrupole mass spectrometry is performed with 0.1% formic acid acetonitrile as mobile phase A and 0.1% formic acid solution as mobile phase B, and the gradient elution is performed: 0-3min, 5%; the flow rate was 0.5ml per minute.
10. The method of claim 5, wherein the triple quadrupole mass spectrometry is monitored by multiple reactions using a mass spectrometer detector in electrospray positive ion mode, and wherein SEQ ID No.1 selects the quantitative ions with mass to charge ratio m/z 639.6 → 810.2, the qualitative ions with mass to charge ratio m/z 639.6 → 753.1 are used as the detecting ion pair, and SEQ ID No.2 selects the quantitative ions with mass to charge ratio m/z852.1 → 1144.6, and the qualitative ions with mass to charge ratio m/z852.1 → 875.4 are used as the detecting ion pair.
CN202210653950.9A 2022-06-10 2022-06-10 Syngnathus characteristic polypeptide, application thereof and method for identifying Syngnathus Pending CN115246873A (en)

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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN111303263A (en) * 2020-03-11 2020-06-19 中国药科大学 Earthworm characteristic polypeptide and application thereof in species identification of earthworm medicinal materials
CN114113381A (en) * 2021-11-12 2022-03-01 山东省食品药品检验研究院 Characteristic polypeptide of saloon, application thereof and method for identifying comfortable saloon

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN111303263A (en) * 2020-03-11 2020-06-19 中国药科大学 Earthworm characteristic polypeptide and application thereof in species identification of earthworm medicinal materials
CN114113381A (en) * 2021-11-12 2022-03-01 山东省食品药品检验研究院 Characteristic polypeptide of saloon, application thereof and method for identifying comfortable saloon

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