Picomolar quantitation of free sulfite in foods by means of [57Co]hydroxocobalamin and radiometric chromatography of [57Co]sulfitocobalamin. Method, applications and significance of coexisting sulfides.

The concentration dependent reaction of sulfite with 57Co-labeled hydroxocobalamin (OH57CoCbl) to produce a sulfitocobalamin (SO(3)57CoCbl) adduct served as a quantification strategy for foodborne sulfite residues freely extracted into pH 5.2, 0.05 M acetate buffer. SO(3)57CoCbl was then resolved using SP-Sephadex C-25 gel chromatography and its radiometric detection allowed calculation of a standard logit plot from which unknown sulfite concentrations could be determined. The sulfite detection range was 6.0 nM-0.3 pM with respective relative standard deviations of 4.4-29.4% for 50-microl samples. Individual incidences of foodborne sulfite intolerances provoked by L-cysteine or sulfite additive use in bakery products, which remained undetected using conventional sulfite analytical methods, underscored the quantitative value of the method. The analytical significance and occurrences of detectable sulfides coexisting with foodborne sulfite residues was also addressed.

Nitritocobalamin and nitrosocobalamin may be confused with sulfitocobalamin using cation-exchange chromatography.

The previously reported ability of SP-Sephadex C25 column chromatography for partitioning biologically important cobalamins has been modified to include analytical separation of nitritocobalamin (NO2-Cbl) and nitrosocobalamin (NO-Cbl). Gel column dimensions (1.5 x 11.0 cm), a low eluent flow-rate (125 microliters/min), collection of small eluate fractions (160 microliters) plus maintenance of He saturated mobile and gel phases all combined to eliminate ordinarily confusing proximal elution of NO2-Cbl and NO-Cb1 with sulfitocobalamin (SO3-Cbl) and cyanocobalamin. Cobalamin elution profiles from the gel column were monitored by direct radiometric analysis of 57Co-labelled cobalamin standards or competitive intrinsic factor radioassays for cobalamin sample sizes up to 10.0 ng. Failure to implement the chromatographic conditions detailed here totally obscured analysis of NO2-Cbl coexisting with SO3-Cbl in brain tissues for chicks exposed to dietary sulfites and caused oversight of NO-Cb1 normally coexisting in prepared NO2-Cbl standards.