A Sensitive Method for Detection of Sulfamethazine and N⁴-Acetylsulfamethazine Residues in Environmental Samples Using Solid Phase Immunoextraction Coupled with MALDI-TOF MS

Abstract

Sulfamethazine (SMT) and its major metabolite, N(4)-acetylsulfamethazine (NA-SMT), were each recovered from spiked water (0.1 ppb) and 10% (w/v) aqueous suspensions of soil (1 ppb) or composted manure (1 ppb), by using a three-stage solid phase immunoextraction (SPIE) system, followed by detection with matrix-assisted laser/desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Sulfonamide recovery rates are reported for separate stages of the SPIE system and for trace-level sulfonamide SPIE extraction from the environmental samples. SPIE MALDI-TOF MS is a rapid and definitive technique with potentially better efficiency relative to other established trace-level sulfonamide analytical methods. SPIE MALDI-TOF MS required 1.5 h per batch (8-24 samples/batch) for sample enrichment, 5 min per batch for probe preparation, and 5 min per sample to acquire and process the spectrum. This is the first time MALDI-TOF MS has been reported as a potential means of detecting trace-level drug residues in complex environmental samples.

Sorption, Fate, and Mobility of Sulfonamides in Soils

“…Nonetheless, many animal and human studies have shown that the metabolites of SMZ include N4-acetylSMZ (NA-SMZ), desamino-SMZ, N4-D-glucosyl-SMZ, N4-(1-deoxy-D-glucuronyl)-SMZ (Shaikh et al 1999), and its two hydroxyl metabolites, 6-hydroxymethylSMZ and 5-hydroxySMZ (Nouws et al 1985). The metabolite, NA-SMZ (Table 1), has been found to be a major metabolite of SMZ, and was frequently detected in water, soil, and manure (Grant et al 2003). The acetyl group of NA-SMZ is readily hydrolyzed (Grant et al 2003) and the ionized NA-SMZ is more polar than the parent SMZ.…”

“…The metabolite, NA-SMZ (Table 1), has been found to be a major metabolite of SMZ, and was frequently detected in water, soil, and manure (Grant et al 2003). The acetyl group of NA-SMZ is readily hydrolyzed (Grant et al 2003) and the ionized NA-SMZ is more polar than the parent SMZ. Also, degradation pathways in the environment, through bacterial or enzymatic processes, may be different from those determined in human and animals.…”

Generic anti-sulfonamide immunoaffinity columns made using sulfamethazine-specific polyclonal antibodies

“…SMT and STZ were also used to make N 4 -azo and N 4 -succinyl conjugates of sulfonamide-Limulus polyphemus hemolymph (LPH) and bovine serum albumin (BSA). The analyses of sulfonamides by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and the colorimetric Bratton-Marshall (BM) assay were described previously (Grant et al 2003).…”

“…They are widely and routinely used in medicine and agriculture, and can persist as residues in food and the environment (Heberer 2002, Kolpin et al 2002. Several analytical techniques, such as enzyme-linked immunosorbent assay (ELISA) (Haasnoot et al 1996, Muldoon et al 2000a, Thomson & Sporns 1995, Haasnoot et al 2000, Spinks et al 2002, Lee et al 2001, Sheth & Sporns 1991 and immunoaffinity chromatography (Crabbe et al 1999, Li et al 2000, Grant et al 2003 rely upon antibodies to recognize sulfonamides. Although immunological techniques usually possess exquisite specificity, cross-reactivity is an inherent property of any antibody molecule toward antigens related to the immunogen.…”

“…When a cross-reactive immunoaffinity column (IAC) is desired, it may be advantageous to use pAbs instead of monoclonal antibodies, since their inherent heterogeneity may offer greater cross-reactivity. Our previous work in which immunoextraction was coupled to mass spectrometry to detect sulfonamide residues in environmental samples (Grant et al 2003), generated a number of questions about the nature of IAC recovery of sulfamethazine and its N 4 -acetyl metabolite. We found that the design of the linking arm used to make the sulfonamide-protein immunogen had a large bearing on antibody specificity to a variety of analytes (Grant & Sporns 2005).…”