Solid phase extraction-high performance liquid chromatography-fluorescence method for the determination of monoamine neurotransmitters and their metabolites in human urine

Abstract

A method for determining monoamine neurotransmitters and metabolites thereof in human urine through solid-phase extraction-high performance liquid chromatography-fluorescence spectrometry includes thefollowing steps: S1, preprocessing a sample; S2, preparing a sample: performing solid phase extraction by using a self-made solid phase extraction column; S3, preparing a standard solution; S4, drawing a standard curve; and S5, determining: introducing the sample obtained in step S2, performing HPLC analysis, qualitatively analyzing the retention time, and quantifying the peak area. Cellulose acetate is used as a carrier, pomelo peel and flesh powder is used as an adsorbent, the pomelo peel and flesh powder is evenly distributed in the gaps of the cellulose acetate, and the peel and flesh powder has excellent sensitivity and selectivity to the monoamine neurotransmitters and the metabolites thereof in the urine, so the monoamine neurotransmitters and the metabolites thereof in the human urine are enriched in pomelo peel and flesh powder, the purity and the sensitivity of the sample are greatly improve, the detection limit is reduced, the recovery rate is improved, and the detection efficiency is improved; and the cost of the method is low, so the detection cost is reduced.

Description

technical field

[0001] The invention relates to the technical field of determination methods for monoamine neurotransmitters and their metabolites in human urine, in particular to the determination of monoamine neurotransmitters and their metabolism in human urine by solid phase extraction-high performance liquid chromatography-fluorescence method way of things. Background technique

[0002] Neurotransmitters are chemical substances secreted by nerve endings that act as messengers in chemical synaptic transmission. Monoamine neurotransmitters (MNTs) were the first class discovered, including dopamine (DA), epinephrine (EP), norepinephrine (NE), and serotonin (5-HT) and its metabolite vanilla almond acid (VMA), homovanillic acid (HVA), 5-hydroxyindoleacetic acid (5-HIAA). MNTs play an extremely important role in the central nervous system, peripheral autonomic nervous system and hormone-mediated endocrine and exocrine activities. Solid-phase extraction (Solid-Phase Extract...

Examples

Embodiment 1

[0029] see figure 1 , the method for solid-phase extraction-high performance liquid chromatography-fluorescence assay monoamine neurotransmitters and their metabolites in human urine, comprising the following steps:

[0030] S1. Pretreatment of samples: take 2ml of fresh human urine, add 200μl of 1mol / L hydrochloric acid solution to acidify the urine, then add 2.0ml of 0.5mol / L ammonium acetate, mix well, ultrasonicate for 5min, and then use 5000r Centrifuge for 10 min at 1 / min, take the supernatant, filter through a 0.45 μm membrane filter, and store the filtrate in a refrigerator at 4°C until use;

[0031] S2. Sample preparation: Dry and pulverize pomelo husk through a 300-mesh sieve to obtain pomelo husk powder, then roast the pomelo husk powder under vacuum conditions for 0.6h at a temperature of 550°C to obtain pomelo husk adsorbent , fill the cellulose acetate in the cigarette filter in the SPE glass tube, then add the pomelo peel adsorbent, and add a gasket to obtain t...

Embodiment 2

[0039] The method for measuring monoamine neurotransmitters and metabolites thereof in human urine by solid phase extraction-high performance liquid chromatography-fluorescence method comprises the following steps:

[0040] S1. Pretreatment of samples: take 2ml of fresh human urine, add 200μl of 1mol / L hydrochloric acid solution to acidify the urine, then add 2.0ml of 0.5mol / L ammonium acetate, mix well, ultrasonicate for 5min, and then use 5000r Centrifuge for 10 min at 1 / min, take the supernatant, filter through a 0.45 μm membrane filter, and store the filtrate in a refrigerator at 4°C until use;

[0041] S2. Preparation of samples: drying and pulverizing the pomelo husk through a 500-mesh sieve to obtain pomelo husk powder, and then roasting the pomelo husk powder under vacuum conditions for 1 hour at a temperature of 300° C. to obtain the pomelo husk adsorbent. Fill the cellulose acetate in the cigarette filter in the SPE glass tube, then add the pomelo peel adsorbent, and...

Embodiment 3

[0049] The method for measuring monoamine neurotransmitters and metabolites thereof in human urine by solid phase extraction-high performance liquid chromatography-fluorescence method comprises the following steps:

[0050]S1. Pretreatment of samples: take 2ml of fresh human urine, add 200μl of 1mol / L hydrochloric acid solution to acidify the urine, then add 2.0ml of 0.5mol / L ammonium acetate, mix well, ultrasonicate for 5min, and then use 5000r Centrifuge for 10 min at 1 / min, take the supernatant, filter through a 0.45 μm membrane filter, and store the filtrate in a refrigerator at 4°C until use;

[0051] S2. Sample preparation: Dry and pulverize pomelo husk through a 400-mesh sieve to obtain pomelo husk powder, then roast pomelo husk powder under vacuum conditions for 0.8 hours at 480°C to obtain pomelo husk adsorbent , fill the cellulose acetate in the cigarette filter in the SPE glass tube, then add the pomelo peel adsorbent, and add a gasket to obtain the self-made solid ...

technical field

[0001] The invention relates to the technical field of determination methods for monoamine neurotransmitters and their metabolites in human urine, in particular to the determination of monoamine neurotransmitters and their metabolism in human urine by solid phase extraction-high performance liquid chromatography-fluorescence method way of things. Background technique

[0002] Neurotransmitters are chemical substances secreted by nerve endings that act as messengers in chemical synaptic transmission. Monoamine neurotransmitters (MNTs) were the first class discovered, including dopamine (DA), epinephrine (EP), norepinephrine (NE), and serotonin (5-HT) and its metabolite vanilla almond acid (VMA), homovanillic acid (HVA), 5-hydroxyindoleacetic acid (5-HIAA). MNTs play an extremely important role in the central nervous system, peripheral autonomic nervous system and hormone-mediated endocrine and exocrine activities. Solid-phase extraction (Solid-Phase Extract...

Examples

Embodiment 1

[0029] see figure 1 , the method for solid-phase extraction-high performance liquid chromatography-fluorescence assay monoamine neurotransmitters and their metabolites in human urine, comprising the following steps:

[0030] S1. Pretreatment of samples: take 2ml of fresh human urine, add 200μl of 1mol / L hydrochloric acid solution to acidify the urine, then add 2.0ml of 0.5mol / L ammonium acetate, mix well, ultrasonicate for 5min, and then use 5000r Centrifuge for 10 min at 1 / min, take the supernatant, filter through a 0.45 μm membrane filter, and store the filtrate in a refrigerator at 4°C until use;

[0031] S2. Sample preparation: Dry and pulverize pomelo husk through a 300-mesh sieve to obtain pomelo husk powder, then roast the pomelo husk powder under vacuum conditions for 0.6h at a temperature of 550°C to obtain pomelo husk adsorbent , fill the cellulose acetate in the cigarette filter in the SPE glass tube, then add the pomelo peel adsorbent, and add a gasket to obtain t...

Embodiment 2

[0039] The method for measuring monoamine neurotransmitters and metabolites thereof in human urine by solid phase extraction-high performance liquid chromatography-fluorescence method comprises the following steps:

[0040] S1. Pretreatment of samples: take 2ml of fresh human urine, add 200μl of 1mol / L hydrochloric acid solution to acidify the urine, then add 2.0ml of 0.5mol / L ammonium acetate, mix well, ultrasonicate for 5min, and then use 5000r Centrifuge for 10 min at 1 / min, take the supernatant, filter through a 0.45 μm membrane filter, and store the filtrate in a refrigerator at 4°C until use;

[0041] S2. Preparation of samples: drying and pulverizing the pomelo husk through a 500-mesh sieve to obtain pomelo husk powder, and then roasting the pomelo husk powder under vacuum conditions for 1 hour at a temperature of 300° C. to obtain the pomelo husk adsorbent. Fill the cellulose acetate in the cigarette filter in the SPE glass tube, then add the pomelo peel adsorbent, and...

Embodiment 3

[0049] The method for measuring monoamine neurotransmitters and metabolites thereof in human urine by solid phase extraction-high performance liquid chromatography-fluorescence method comprises the following steps:

[0050]S1. Pretreatment of samples: take 2ml of fresh human urine, add 200μl of 1mol / L hydrochloric acid solution to acidify the urine, then add 2.0ml of 0.5mol / L ammonium acetate, mix well, ultrasonicate for 5min, and then use 5000r Centrifuge for 10 min at 1 / min, take the supernatant, filter through a 0.45 μm membrane filter, and store the filtrate in a refrigerator at 4°C until use;

[0051] S2. Sample preparation: Dry and pulverize pomelo husk through a 400-mesh sieve to obtain pomelo husk powder, then roast pomelo husk powder under vacuum conditions for 0.8 hours at 480°C to obtain pomelo husk adsorbent , fill the cellulose acetate in the cigarette filter in the SPE glass tube, then add the pomelo peel adsorbent, and add a gasket to obtain the self-made solid ...

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