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

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.

1 . A method for determining the presence of one or more analytes in a test sample, the method comprising:
 a) extraction and purification of the one or more analytes from the test sample with one or more of solid phase extraction, supported liquid extraction (SLP), and liquid liquid extraction (LLP);   b) derivatization of the one or more analytes with a crown-ether derivatizing agent;   c) detection of the one or more derivatized analytes using liquid chromatography and/or mass spectrometry.   
     
     
         2 . The method of  claim 1 , wherein the one or more analytes is a drug, a hormone, a signaling agent, an amino acid, or a pesticide. 
     
     
         3 . The method of  claim 1 , wherein the one or more analytes is a monoamine neurotransmitter including vitamin D or one of its derivatives or metabolites, a sex hormone or one of its derivatives or metabolites, a cannabinoid or one of its derivatives or metabolites, an opiate, opioid or one of its derivatives or metabolites or an arylcyclohexylamine or one of its derivatives or metabolites, an Amphetamine or one of its derivatives or metabolites 
     
     
         4 . The method of  claim 3 , wherein the monoamine neurotransmitter is Histamine, Tryptamine, Serotonin, or Agmatine; wherein the sex hormone or one of its derivatives or metabolites is an estrogen; wherein the derivative of vitamin D is 25-OH D 3 , 25-OH D 2 , 24,25-(OH) 2  D 3 , 1,25-(OH) 2  D 3 , and 1,25-(OH) 2  D 2 , Cholecalciferol, 25-Hydroxycholecalciferol, 1α,25-Dihydroxycholecalciferol, Ergocalciferol, 1α,25-Dihydroxyergocalciferol, 22,23-Dihydroergocalciferol, 1α,24R,25-Trihydroxycholecalciferol, (6Z)-tacalciol, Tachysterol 3 , Isovitamin D 3 , Dihydrotachysterol 3 ; wherein the cannabinoid or one of its derivatives or metabolites is a Cannabigerol-type (CBG) cannabinoid, a Cannabichromene-type (CBC) cannabinoid, a Cannabidiol-type (CBD) cannabinoid, a Cannabinodiol-type (CBND) cannabinoid, a Tetrahydrocannabinol-type (THC) cannabinoid, a Cannabinol-type (CBN) cannabinoid, a Cannabitriol-type (CBT) cannabinoid, a Cannabielsoin-type (CBE) cannabinoid, an Isocannabinoid, a Cannabicyclol-type (CBL) cannabinoid, a Cannabicitran-type (CBT) cannabinoid, or a Cannabichromanone-type (CBCN) cannabinoid; wherein the opiate, the opioid or the derivative or metabolite of the opiate or opioid, is morphine, oripavine, morphinone, hydromorphone, oxymorphone, a benzylisoquinoline alkaloid, a semi-synthetic or a benzylisoquinoline alkaloid derivative; wherein the arylcyclohexylamine or one of its derivatives or metabolites is Tiletamine, 3-Methoxetamine (MXE), Methoxyketamine, N-Ethylnorletamine (Ethketamine), Amphetamine, Ephedrine, or Methamphetamine; or wherein the Amphetamine or one of its derivatives or metabolites is Amphetamine (itself), methamphetamine, ephedrine, cathinone, 3,4-methylenedioxy-N-methylamphetamine (MDMA, “Ecstasy”), and 2,5-Dimethoxy-4-methylamphetamine (DOM, or “STP”). 
     
     
         5 . The method of  claim 1 , wherein the solid phase extraction is performed with an ion exchange column or cartridge, wherein the ion exchange column is a cation exchange column, a weak cation exchange column or an anion exchange column. 
     
     
         6 . The method of  claim 1 , wherein the solid phase extraction is performed with a reverse phase silica column or cartridge. 
     
     
         7 . The method of  claim 6 , wherein the reverse phase silica is an alkyl bounded (C4, C8, C12, or C18) silica, a cyano bounded silica, a phenyl bounded silica, or a biphenyl bounded silica. 
     
     
         8 . The method of  claim 1 , wherein the sample is a biological sample, a soil sample, or a sample of food stuff. 
     
     
         9 . The method of  claim 8 , wherein the biological sample is a blood sample, a saliva sample, a lachrymal sample, a urine sample, or a tissue sample. 
     
     
         10 . The method of  claim 9 , wherein the blood sample is a full blood sample, a plasma sample, or a serum sample. 
     
     
         11 . The method of  claim 1 , wherein the crown-ether derivatizing agent comprises: a crown-ether, a connector, and an analyte-binding functional group. 
     
     
         12 . The method of  claim 11 , wherein the crown-ether comprises a ring, and the ring is a 12-30 membered ring. 
     
     
         13 . The method of  claim 12 , wherein the ring has 12-30 member atoms of which 8-20 atoms are carbon and wherein the non-carbon member atoms are selected from oxygen, nitrogen, and sulfur. 
     
     
         14 . The method of  claim 11 , wherein the crown ether is selected from 12 Crown 4, 15 Crown 5, 16 crown 4, 18 Crown 6, 21 Crown 7, or 24 Crown 8, optionally having one or more heteroatoms replacing oxygen. 
     
     
         15 . The method of  claim 11 , wherein the connector is a C 1 -C 12  linear, branched, and/or cyclic alkyl group or a phenolic ring fused to the crown ether. 
     
     
         16 . The method of  claim 11 , wherein the analyte binding group is an acylating group, 4-Phenyl-1,2,4-triazolin-3,5-dione (PTAD), 1,2,4-traizoline-3,5-dione (TAD), an Alkoxylamine, a hydrazide, an alcohol, or an amine. 
     
     
         17 . The method of  claim 16 , wherein the acylating group is an acylating agent of Formula 2: 
       
         
           
           
               
               
           
         
         where A is the analyte and X is the connector. 
       
     
     
         18 . The method of  claim 17 , wherein the acylating agent that is an acyl chloride or acyl halide. 
     
     
         19 . The method of  claim 1 , where the crown ether derivatizing agent is selected from the agents of Group I, II, III, or IV: 
       
         
           
           
               
               
           
         
         
           
           
               
               
           
         
         
           
           
               
               
           
         
         
           
           
               
               
           
         
         
           
           
               
               
           
         
         
           
           
               
               
           
         
         
           
           
               
               
           
         
         
           
           
               
               
           
         
         
           
           
               
               
           
         
       
     
     
         20 . The method of  claim 19 , wherein the derivatizing agent is MB338 or MB409. 
     
     
         21 . A derivatization reagent for mass spectrometry having a derivatizing agent comprising: a crown-ether, a connector, and an analyte-binding functional group.