High-pressure liquid chromatography and field desorption mass spectrometry of heme a, Heme a dimethyl ester and acetyl heme a dimethyl ester

Abstract

Field desorption mass spectrometry was shown to be a valid technique for determining the molecular weights of hemins and hemin esters, as well as of porphyrins. The observed base peaks of ligand-free protoheme IX, protoheme IX dimethyl ester, and protoporphyrin IX dimethyl ester correspond well to the molecular weights of these compounds and the base peak for hematoporphyrin corresponds to the molecular weight of this porphyrin minus two molecules of water. The technique was employed to confirm the molecular weights of heme a, heme a dimethyl ester, and acetyl heme a dimethyl ester. Heme a dimethyl ester was prepared by reaction of heine a with trimethyloxonium tetrafluoroborate and purified by high-pressure liquid chromatography. The isolated product was converted to the acetylated derivative by reaction with acetic anhydride and was subsequently purified by high-pressure liquid chromatography. A field desorption spectrum of heme a shows a base peak at 582 which is in agreement with previous deductions of the structure of this prosthetic group. Base peaks of the heme a ester and its acetylated derivative demonstrate that the two carboxyl groups have been methylated and the single hydroxyl group has been acetylated without further alteration of the molecule.

Reactivities of hydroxylamine and sodium bisulphite with carbonyl-containing haems with the prosthetic groups of the erythrocyte green haemoproteins

“…The haemochrome of haem a was prepared in pyridine/water (9: 1, v/v) rather than in pure pyridine. As reported previously (DeFilippi & Hultquist, 1977), the spectra in pyridine/water and in pure pyridine are identical. An absorption coefficient of 32.7 mm-' cm-' (Caughey et al, 1975) was used for the a-peak of haem a.…”

“…This perturbation may be a result of simple electron-withdrawing ability. Such an explanation is weakened, however, by the failure of the a-hydroxyfarnesylethyl group to act as a rhodo-1979 fying group in the porphyrin spectrum (Lemberg & Falk, 1951) and the failure of acetylation of the hydroxy group to perturb the reduced pyridine haemochrome spectrum of haem a dimethyl ester (DeFilippi & Hultquist, 1977). Alternatively, the perturbation may result from affects on the solvation of the porphyrin system, or, most likely, from interaction of the double bonds of this side chain with the porphyrin 7-electron system as has been suggested by Caughey et al (1975).…”

Comparison of the sensitivity of mass spectrometry atmospheric pressure ionization techniques in the analysis of porphyrinoids

“…Porphyrinoids are characterized by a great variety of structure types; hence, it is not possible to analyze them by using only one ionization method. There are many examples in the literature regarding the application of various MS ionization techniques [EI, [2,3] chemical ionization (CI), [4,5] electrospray ionization (ESI), [6][7][8][9][10][11] atmospheric pressure CI (APCI), [12][13][14] atmospheric pressure photoionization (APPI), [15][16][17][18] fast atom bombardment (FAB), [19][20][21] liquid secondary ion MS, [22,23] field desorption, [24][25][26] matrix-assisted laser desorption/ionization (MALDI) [19,27,28] ] with reference to this class of compounds. It is also possible to find examples of comparison of ionization methods in qualitative terms, but the number of such publications is very limited.…”