Method for in-situ analysis of squalene content in microbial colonies

A microbial colony and in-situ analysis technology, applied in the field of analysis and detection, can solve problems such as inability to achieve high-throughput in-situ analysis, and achieve the effect of reducing acquisition time and saving time

Active Publication Date: 2021-08-17
QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these techniques still require sample preparation, matrix doping, or extraction with strong organic reagents, which cannot achieve high-throughput in situ analysis of the original colony and its surrounding environment

Method used

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  • Method for in-situ analysis of squalene content in microbial colonies
  • Method for in-situ analysis of squalene content in microbial colonies
  • Method for in-situ analysis of squalene content in microbial colonies

Examples

Experimental program
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Effect test

Embodiment 1

[0032] A laser electrospray mass spectrometry device for in-situ analysis of a single colony of microorganisms, including a laser 1, an electrospray device 2, a displacement platform 3 and a mass spectrometer 4, the laser 1 is located directly above the displacement platform 3, and the electrospray device 2 is located at the displacement platform Above the front of the platform 3 , the mass spectrometer 4 is located above the rear of the displacement platform 3 , and the electrospray device 2 is located directly in front of the mass spectrometer 4 . A sample plate 27 for cultivating the target bacterial strain is arranged on the displacement plate 3 . The electrospray device 2 is fixed on the round hole bracket 18; the round hole bracket 18 is fixed on the three-dimensional micro-adjustment base 19, and the front and rear micro-adjustment knobs 20, left and right micro-adjustment Knob 21 and fine adjustment knob 22 up and down. The displacement platform 3 is fixed on the hori...

Embodiment 2

[0035] Embodiment 2: Determination of squalene content in the single colony of marine yeast cultured on the plate

[0036] The agar plate coated with 50 μL of marine yeast liquid was cultured in a 25°C incubator for 24 hours, and a total of 186 single colonies grew out, with a maximum diameter of about 10 mm and an average diameter of about 5 mm; place the plate 15 mm below the laser interface At , set the Nd:YAG solid-state pulsed laser with an energy of 600mJ / cm 2 , the formed laser spot is about 0.5mm. The gas-phase molecular plume of metabolites formed after the laser bombards the colony cells and the external electrospray interface (the voltage applied by the front-end quartz electrospray needle is 5kV, and the inner diameter of the quartz electrospray needle is 50 μm; the outlet of the quartz electrospray needle The inner diameter of the tip is 5 μm; the injection flow rate of the solvent reagent added into the injection needle is 100nL / min), and the high-voltage charge...

Embodiment 4

[0045] Embodiment 4: the detection of squalene content in the chlorella single algal colony cell on the plate

[0046] Incubate the agar plate coated with 80 μL of Schizochytrium bacteria solution in a 33°C incubator for 18 hours, and a total of 173 single colonies grew out, with a maximum diameter of about 8 mm and an average diameter of about 4 mm; place the plate on the laser interface At 10mm below, set the Nd:YAG solid-state pulse laser with an energy of 400mJ / cm 2 , the formed laser spot is about 1mm. The gas-phase molecular plume of metabolites formed after the laser bombards the colony cells and the external electrospray interface (the voltage applied by the front-end quartz electrospray needle is 4kV; the inner diameter of the quartz electrospray needle is 60 μm; the outlet of the quartz electrospray needle The inner diameter of the tip is 15 μm; the injection flow rate of the solvent reagent added into the injection needle is 400nL / min), and the high-voltage charged...

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Abstract

Aiming at the problems existing in the detection of squalene content in high-throughput screening of squalene-producing strains in the prior art, the invention provides a method for in situ analysis of squalene content in microbial colonies. The method for in-situ analysis of squalene content in microbial colonies of the present invention includes (1) sample preparation (cultivating microbial colonies to a suitable size on an agar plate), (2) laser bombardment ablation and desorption of single colony cells Formation of gas-phase molecular plume, (3) high-pressure charged mist droplets produced by the electrospray interface interact with gas-phase molecular plume to charge metabolites, (4) charged metabolites are inhaled into mass spectrometer for detection and (5) external standard method to calculate content 5 steps. The method of the present invention is based on laser ablation electrospray ionization mass spectrometry technology, realizes the in-situ detection of squalene in the bacterial colonies on the plate, and the detection of the extraction-free preparation process, thereby realizing high-throughput rapid analysis and filling the existing technology It overcomes the technical bottleneck of screening high-yield squalene strains in the prior art.

Description

technical field [0001] The invention belongs to the field of analysis and detection, in particular to a detection method for squalene content, in particular to a method for in-situ analysis of squalene content in microbial colonies. Background technique [0002] Squalene (squalene, 2,6,10,15,19,23-hexamethyl-2,6,10,14,18,22-tetracosahexaene) belongs to open-chain triterpenes, which have the function of enhancing body Immunity, improvement of sexual function, anti-aging, anti-fatigue, anti-tumor and other physiological functions are widely used in medicine, beauty, cosmetics and other fields. Although squalene widely exists in animals, plants and microorganisms, at present, the liver oil of deep-sea sharks is still the main source of squalene. With the depletion of fishery resources and the improvement of related laws and regulations, people are trying to find other sources of squalene. Possible sources of squalene include extraction and isolation from olive oil, palm oil, ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N27/64
CPCG01N27/64
Inventor 宋晓金刘欢崔球王森张慧丹万伟建蓝传增
Owner QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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