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Solid Phase Extraction, Derivatization with Crown Ethers, and Mass Spectrometry, Methods, Reagents and Kits

a technology of derivatization and mass spectrometry, which is applied in the field of solid phase extraction, derivatization with crown ethers, mass spectrometry, methods, reagents and kits, can solve the problems of complex and cumbersome extraction procedures for quantification of certain analytes, and difficult detection of certain analytes, etc., to achieve similar sensitivity and accuracy, simple process, and easy configuration

Inactive Publication Date: 2016-09-29
TECAN SP INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

The patent describes a new method for measuring small amounts of substances in blood plasma or oral fluid using a combination of solid phase extraction and chemical derivatization with a specific reagent. This method is sensitive and accurate, with a minimum detection limit of picograms per milliliter, and can be easily automated. Unlike other methods, this technique does not produce analyte cross-reactivity, which makes it a more robust and reliable tool for clinical settings.

Problems solved by technology

However, certain analytes may be difficult to detect for a multitude of reasons.
For instance, some analytes are difficult to detect as they are prone to be oxidized and degraded.
Further, the current art in quantification of certain analytes often involves complex and cumbersome extraction procedures with unstable extraction recovery.
For instance, unstable extraction recovery gives rise to results which are often unreliable, and with high lower limits of quantification (LLOQ); for example an LLOQ in >10 ng / mL range.
Also, the existing extraction process is unable to provide a reliable sample for derivatization.
Again, these drugs may not be stable in bodily fluids and therefore, detection may be difficult.

Method used

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  • Solid Phase Extraction, Derivatization with Crown Ethers, and Mass Spectrometry, Methods, Reagents and Kits
  • Solid Phase Extraction, Derivatization with Crown Ethers, and Mass Spectrometry, Methods, Reagents and Kits
  • Solid Phase Extraction, Derivatization with Crown Ethers, and Mass Spectrometry, Methods, Reagents and Kits

Examples

Experimental program
Comparison scheme
Effect test

example 1

Quantification of 1,25-Dihydroxy-Vitamin D in Plasma

Sample Collection

[0207]Plasma samples were obtained from human patients' blood. Samples were drawn (plasma sodium heparin & EDTA) into pre-chilled Vacutainers. Vacutainers were inverted 5× and refrigerated until centrifuged. Plasma was separated in a refrigerated centrifuge (1000×g for 10 minutes) within 30 minutes of collection and then frozen immediately at −20° C. in plastic vials.

[0208]Plasma was thawed and diluted before use in solid phase extraction.

Standards

[0209]The blanks, calibration samples, and plasma samples were spiked with internal standards (e.g., D6-1,25(OH)2 vitamin D3 and / or D6-1,25(OH)2 vitamin D2).

[0210]Standard curves were generated with plasma solutions spiked with a known amount of 1,24-dihydroxy-vitamin D. The spiking solution was serially diluted before being added to the plasma taken from the same plasma sample.

Extraction

[0211]TRACE-N® (3 cc / 15 mg) columns were conditioned with 1.0 ml of methanol, followe...

example 2

Quantification of THC or HU210 in Oral Fluid

Sample Collection

[0222]Oral samples were taken from humans using the QUANTISAL™ (Immunalysis, California). Saliva was taken from patients using the collection device from QUANTISAL, and specimens were refrigerated and ultimately frozen. Saliva was thawed before use in the present quantification methods.

Standards

[0223]The blanks, calibration samples, and oral samples were spiked with internal standards (e.g., THC-d3).

[0224]Standard curves were generated with saliva solutions spiked with a known amount of THC or HU210 (a synthetic cannabinoid). The spiking solution was serially diluted before being added to the saliva taken from the same oral sample.

Extraction

[0225]TRACE-N® (3 cc / 15 mg) columns were conditioned with 0.5 ml of methanol, followed by 0.5 ml 0.1% HCl (6M) in water.

[0226]Then 300 uL of the oral fluid in QUANTISAL™ buffer (25% v / v). 10 μL internal standard was added and 600 uL of 100 mM pH6 phosphate buffer. The Sample / buffer mix ...

example 3

Quantification of Estradiol / Estriol / Estrone / 17β-Estradiol-2,3,4-13C3 in Plasma

Sample Collection

[0235]Plasma samples were obtained from human patients' blood. Samples were drawn (plasma sodium heparin & EDTA) into pre-chilled Vacutainers. Vacutainers were inverted 5× and refrigerated until centrifuged. Plasma was separated in a refrigerated centrifuge (1000×g for 10 minutes) within 30 minutes of collection and then frozen immediately at −20° C. in plastic vials.

[0236]Plasma was thawed and diluted before use in solid phase extraction.

Standards

[0237]The blanks, calibration samples, and plasma samples were spiked with internal standards (e.g., 17β-estradiol-2,3,4-13C3).

[0238]Standard curves were generated with plasma solutions spiked with a known amount of estradiol or estrone. The spiking solution was serially diluted before being added to the plasma taken from the same plasma sample.

Extraction

[0239]MAESTRO® A (1 cc / 15 mg) columns were conditioned with 1.0 mL of methanol, followed by 0...

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Abstract

The present disclosure is directed to methods reagents and kits for solid phase extraction, derivatization with crown ether containing derivatizing agents, and mass spectrometry of the derivatized analytes.

Description

PRIORITY[0001]This patent application claims the right of priority pursuant to 35 U.S.C. §119(e) and is entitled to the benefit of the filing date of U.S. Provisional Patent Application Ser. No. 62 / 139,692, filed on Mar. 28, 2015, the content of which is hereby expressly incorporated by reference in its entirety.INTRODUCTION[0002]There is a need to obtain fast, streamlined, and automated methods for detection of particular analytes. In the medical context, such methods would facilitate the detection of analytes, such as drugs, hormones, signaling agents, and amino acids. In the agricultural and public health context, the detection of residual levels of antibiotics, pesticides or other contaminants is integral to the safety of the food we eat and water we drink. Furthermore, recent changes in the standards for food labeling, such as “hormone free” or “organic” have created a need for streamlined testing in the agricultural realm.[0003]In many cases clinical quantification of particul...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): G01N33/94G01N33/82
CPCG01N33/948G01N2560/00G01N33/82G01N33/743C07K1/36C07K1/00G01N33/6848
Inventor HUANG, QIDIMSON, PHILIPMALOLES CHANCO, EMMANUEL LUIS
Owner TECAN SP INC
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