Ultrasonic-assisted pretreatment method for extraction of multiple steroid hormones in sediment

Patent metadata

Field	Value
Publication number	US-11976093-B2
Application number	US-202117170969-A
Country	US
Kind code	B2
Filing date	Feb 9, 2021
Priority date	May 5, 2019
Publication date	May 7, 2024
Grant date	May 7, 2024

Abstract

The disclosure discloses an ultrasonic-assisted pretreatment method for extraction of multiple steroid hormones in a sediment, including the following steps: (1) lyophilizing the sediment, grinding the sediment, and passing the ground sediment through a 40-60-mesh sieve; (2) placing the sample obtained in step (1) in a container; (3) adding an extractant to the container in step (2), shaking the mixture for 15 s-30 s, centrifuging the mixture to collect an supernatant after ultrasonication, and repeating extraction three times; where the extractants used in the three times of extraction are two of methanol, acetonitrile and acetone; and (4) after mixing the supernatants of the three times of extraction obtained in step (3), concentrating the mixture under a nitrogen flow at 20-30° C., passing the concentrated mixture through a filter, and performing detection. By using the method of the disclosure, the maximum recovery can be up to 100%. The application range is wide, and a recovery of the plurality of steroid hormones each can be up to 73% or above.

First claim

What is claimed is:

1. A pretreatment method for extraction of multiple steroid hormones in a sediment, comprising the following steps:
(1) lyophilizing the sediment, and grinding the sediment to 40-60 mesh;
(2) placing a sample obtained in step (1) in a container;
(3) adding an extractant to the container in step (2) for extraction, shaking a mixture for 15-30 s, ultrasonicating the mixture, centrifuging the mixture to collect a supernatant after ultrasonication, and repeating the extraction three times; and
(4) after mixing the supernatants of the three times of extraction obtained in step (3), concentrating the mixture under a nitrogen flow at 20-30° C., and passing the concentrated mixture through a filter;
wherein in the step (1), the lyophilizing is to pre-freeze the sediment at −60 to −80° C. overnight and lyophilize the sediment at −45° C.;
wherein solvents for the three times of extraction are methanol, acetonitrile and acetonitrile, respectively, or methanol, methanol and acetone, respectively, or methanol, acetone and acetone, respectively; and the pretreatment method does not need an SPE cleaning step.

2. The pretreatment method according to claim 1 , wherein the steroid hormones comprise estrogens, androgens, and progestogens.

3. The pretreatment method according to claim 1 , wherein in the step (2), an amount of the sample placed in the container is 0.5-5.0 g.

4. The pretreatment method according to claim 1 , wherein in the step (2), the container is made of a polytetrafluoroethylene material.

5. The pretreatment method according to claim 3 , wherein in the step (3), a volume of the extractant is 5-10 mL.

6. The pretreatment method according to claim 1 , wherein in the step (3), a frequency of ultrasound in the ultrasonication is 45-60 kHz.

7. A method for detecting a content of steroid hormones in a sediment or sandy soil, comprising: treating a sample by using the pretreatment method according to claim 1 to extract the steroid hormones in the sample; and then analyzing the sample obtained by the extraction by high performance liquid chromatography-tandem mass spectrometry to quantitatively detect the content of the steroid hormones in the sample, wherein the steroid hormones comprise estrogens, androgens and progestogens; and the sample is the sediment or sandy soil.

8. The method according to claim 7 , wherein conditions for the liquid chromatography when analyzing the estrogens are that: a mobile phase A is 0.05% ammonia water, a mobile phase B is acetonitrile, a flow rate is 0.2 mL/min, an injection volume is 5 μL, and a column temperature is 40° C.; and conditions for elution are that: at 0-0.25 min, a mobile phase is 70% mobile phase A +30% mobile phase B; at 0.25-3 min, the mobile phase B is gradually increased to 90%, and a condition is held for 2 min; and then, a column is immediately restored to an initial mobile phase, namely 70% mobile phase A +30% mobile phase B, and a condition is held for 1 min.

9. The method according to claim 8 , wherein conditions for the mass spectrometry when analyzing the estrogens are that: electrospray ionization is used in negative ion mode (ESI − ); a scanning mode is multiselected reaction monitoring (MRM); a capillary voltage is 2.5 kV; a cone voltage and an extractor voltage are 30.0 V and 3.0 V, respectively; an ion source temperature and a desolvation temperature are 150° C. and 350° C., respectively; a cone gas flow and a desolvation gas flow are 50 L/hr and 700 L/hr, respectively; and a collision gas flow is 0.16 mL/min.

10. The method according to claim 7 , wherein conditions for the chromatography when analyzing the androgens and the progestogens are that: a mobile phase A is a methanol solution containing 0.1% of formic acid, a volume ratio of water to methanol in the methanol solution is 98:2, a mobile phase B is acetonitrile, a flow rate is 0.4 mL/min, an injection volume is 5 μL, and a column temperature is 40° C.; and conditions for elution are that: at 0-0.25 min, a mobile phase is 90% mobile phase A +10% mobile phase B; at 0.25-1 min, the mobile phase B is gradually increased to 70%; at 1-3 min, the mobile phase B is increased to 95%, and a condition is held for 1 min; and then, a column is immediately restored to an initial mobile phase, namely 90% mobile phase A +10% mobile phase B, and a condition is held for 1 min.

11. The method according to claim 10 , wherein when detecting the androgens and the progestogens, electrospray ionization is used in positive ion mode (ESI + ), and a scanning mode is multiselected reaction monitoring (MRM).

12. The method according to claim 11 , wherein conditions for the mass spectrometry are that: a capillary voltage is 3.0 kV; a cone voltage and an extractor voltage are 30.0 V and 3.0 V, respectively; an ion source temperature and a desolvation temperature are 150° C. and 500° C., respectively; a cone gas flow and a desolvation gas flow are 50 L/hr and 900 L/hr, respectively; and a collision gas flow is 0.16 mL/min.

13. The method according to claim 7 , wherein the steroid hormones comprise one or more of estrone, estradiol, ethinylestradiol, androstenedione, testosterone, methyltestosterone, levonorgestrel and progesterone.

Assignees

• Univ Jiangnan

Inventors

• Zou Hua
• Luo Xin
• Dai Ruihua
• Zhang Yun
• Shu Shu
• Zhou Zhengkai

Classifications

• C07J75/00 Primary

  Processes for the preparation of steroids in general · CPC title

• B01D11/0257

  using mixing mechanisms, e.g. stirrers, jets (B01D11/0242 takes precedence) · CPC title

• B01D11/0265 Primary

  Applying ultrasound · CPC title

• B01D11/028

  Flow sheets · CPC title

• B01D11/0284 Primary

  Multistage extraction · CPC title