Derivatization Reagents for LC-MS

Introduction

The development of sensitive and selective determination methods in complex matrices for trace level compounds is essential to many research fields. Liquid chromatography-mass spectrometry (LC-MS) has frequently been utilized for this purpose due to its inherent selectivity and sensitivity. However, many compounds cannot be analyzed well by LC-MS, especially if they are difficult to ionize or to fragment, as that makes detection sensitivity extremely low. One of powerful strategies to improve the detection characteristics of compounds in LC-MS is chemical derivatization. And thus a considerable number of derivatization reagents have been synthesized and developed. It aims to modify the structure of the target compounds and, as a consequence, the chemical and physical properties, making the target compound easy to isolate and detect. The general reaction mechanism between target compound and derivatization reagent and the advantages of derivatization-based LC-MS methods are shown in Fig.1 ^[1]. Due to rapid expansion of derivatization-based LC-MS studies, a summary of synthesized derivatization reagent is valuable. Here, Alfa Chemistry provides a comprehensive summary of derivatization reagents for LC-MS studies ^[1-2].

Fig. 1. The general reaction mechanism between target compound and derivatization reagent and the advantages of derivatization-based LC-MS methods.

Summary of Derivatization Reagents

Derivatization reagents can react with target compounds that contain various functional groups, including carbonyl, hydroxyl, carboxyl, amine, thiol, etc. Therefore, it can be categorized into different categories based on the reactions with various functional groups of target compounds. Based on this classification method, we summarize the derivatization reagents for LC-MS.

For Ketones and Aldehydes: The ionization efficiencies of aldehydes or ketones are usually very low, so a chargeable moiety can be introduced to enhance them. The derivatization reagents for this purpose include cyclohexanedione, Girard's reagent P, Girard's reagent T, 2,4-dinitrophenylhydrazine (DNPH), (4-hydrazino-4-oxobutyl) [tris (2,4,6-trimethoxyphenyl) phosphonium] bromide (TMPP-PrG), 5,5'-dimethyl-1,3-cyclohexanedione (Dimethyl CHD), ethoxyamine, 2-hydrazino-1- methylpyridine (HMP), hydroxylamine, 4-hydrazino-N,N,N-trimethyl-4-oxobutanaminium iodide (HTMOB), 2-hydrazinopyridine, 4-[2-(N,N-dimethylamino)ethylaminosulfonyl]-7-N-methylhydrazino-2,1,3-benzoxadiazole (DAABD-MHz), 4-[2-(trimethylammonio)ethoxy] benzenaminium halide (4-APC), 4-{2-[(4-bromophenethyl)dimethylammonio]ethoxy}benzenaminium dibromide (4-APEBA), o-phenylenediamine, D-cysteine, quaternary aminooxy reagent, etc.

Fig. 2. Reactions of (left) HMP and (right) DAABD-MHz with ketones and aldehydes.

For Alcohols and Phenols: Both alcohols and phenols are neutral compounds, so, to improve the MS response, derivatization has been widely employed to in crease the ionization efficiency. The derivatization reagents for this purpose include dansyl chloride (Dns-Cl), dabsyl chloride (DABS-Cl), pyridine-3-sulfonyl chloride (PS), 1,2-dimethylimidazole-4-sulfonyl chloride (DMISC), ethyl chloroformate, benzoyl chloride, 2-sulfobenzoic anhydride, 3-nitrophtalic anhydride, [3-(2-Nitro-4- trifluoromethylphenyl)aminophenyl]dihydroxyborane (2NFP-APB), picolinic acid, 2-bromopyridine-5-boronic acid (BPBA) and m-aminophenylboronic acid (m-APBA), 5-hydroxymethylcytosine, N-methyltrimethylsilyltrifluoroacetamide (MSTFA), S-pentafluorophenyl tris(2,4,6-trimethoxyphenyl)phosphonium acetate bromide (TMPP-AcPFP), 4-(dimethylamino)-benzoic acid (DMBA), diacetyl-L-tartaric anhydride (DATAAN), pentafluorobenzyl bromide (PFBBr), (1-fluoro-2,4- dinitrophenyl-5)-L-alanine amide (L-FDAA), diphenic anhydride, UDP-glucose, etc.

Fig. 3. Reaction of Dns-Cl with alcohols.

For Carboxylic Acids: In the negative ion mode, the carboxylic acid can be analyzed by mass spectrometry. However, the detection sensitivity is usually low. In addition, the mobile phase used for carboxylic acid separation is not always compatible with ESI-MS, so carboxylic acids are often derivatized prior to analysis in order to improve the MS response. The derivatization reagents for this purpose include 3-nitrophenylhydrazine (3-NPH), 2-nitrophenylhydrazine (2-NPH), 4-[2-(N,N-dimethylamino)ethylaminosulfonyl]-7-(2-amino pentylamino)-2,1,3-benzoxadiazole (DAABD-AP), 4-[2-(N,N-dimethylamino)ethylaminosulfonyl]-7-(2-aminobutyl-amino)-2,1,3-benzoxadiazole (DAABD-AB), 4-[2-(N,N-dimethylamino)ethylaminosulfonyl]-7-(2-aminoethylamino)-2,1,3-benzoxadiazole (DAABD-AE), 2-picolylamine, Heptadecafluoroundecylamine (HFUA), 1-pyrenemethylamine (PMA), 1-pyrenemethylamine (PMA), N-[4-(Aminomethyl)phenyl]pyridinium (AMPP), isobutyl chloroformate (i-BuCF), 9-fluorenylmethyl chloroformate (FMOC-Cl), N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide (EDC), p-bromophenacyl bromide (BPBr), 3-bromoactonyltrimethylammonium bromide (BTA), ω-bromoacetonylpyridinium bromide (BPB), etc.

Fig. 4. Reaction of DAABD-AE with carboxylic acids.

For Amines: Challenges for the analysis of amines by LC-MS include their high polarity, basicity and high water solubility. Conversion of amines to more hydrophobic compounds by derivatization is advisable method to solve the problems. The derivatization reagents for this purpose include acetaldehyde, Dns-Cl, succinimidylferrocenyl propionate (SFP), 7-fluoro-4-nitrobenzoxadiazole (NBD-F), 1,2-naphthoquinone-4-sulfonate (NQS), FMOC-Cl, 1,2-benzo-3,4- dihydrocarbazole-9-ethyl chloroformate (BCEOC), hexylchloroformate, 4-nitrobenzylchloroformate, i-BuCF, ethyl-chloroformate, (5-nsuccinimidoxy-5-oxopentyl)triphenylphosphonium bromide (SPTPP), 3-aminopyridyl-N- hydroxysuccinimidyl carbamate (APDS) and the N-hydroxysuccinimide ester of N-alkylnicotinic acid (C_n-NA-NHS), diethyl ethoxymethylenemalonate (DEEMM), N^α-(5-fluoro-2,4-dinitrophenyl)-D-leucinamide (D-FDLA), 7-fluorobenzo-2-oxa-1,3-diazole-4-sulfonate (SBD-F), 4-(dimethylaminoethylaminosulfonyl)-7-chloro-2,1,3-benzoxadiazole (DAABD-Cl), phenyl iso-thiocyanate, methyl acetimidate, phenyl isocyanate, etc.

Fig. 5. Reaction of APDS with amines.

For Thiols: The main problem of thiols analysis is the instability caused by the sample being easily oxidized during processing. Therefore, it is necessary to stabilize these thiol-containing compounds by derivatization during sample preparation. The derivatization reagents for this purpose include Ellman's reagent, 2-bromo-4′-chloroacetophenone (p-CPB), 2-bromo-4′-bromoacetophenone (p-BPB), N-(2-ferroceneethyl)maleimide (FEM), ferrocenecarboxylic acid-(2-maleimidoyl)ethylamide (FMEA), iodoacetamide (IAM), isopropylchloroformate (IPCF), 2,3,4,6-tetra-O-acetyl-β-glucopyranosyl isothiocyanate (GITC), ω-bromoacetonylquinolinium bromide (BQB), Ebselen, etc.

Alfa Chemistry is a global leading supplier of analytical chemistry reagents. We provide a wide range of high quality derivatization reagents for LC-MS studies, and many other analytical chemistry products. If you cannot find the suitable products, Alfa Chemistry also offers you with custom synthesis service. If necessary, please don't hesitate to contact us.

References

Qi B L., et al. Derivatization for liquid chromatography-mass spectrometry[J]. TrAC Trends in Analytical Chemistry, 2014, 59: 121-132.
Santa T. Derivatization reagents in liquid chromatography/electrospray ionization tandem mass spectrometry[J]. Biomedical Chromatography, 2011, 25(1‐2): 1-10.

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