Analyses of Biologically Active Steroids: Antitumor Active OSW-1 and Cardiotonic Marinobufotoxin, by Matrix-Assisted Laser Desorption/Ionization Quadrupole Ion Trap Time-of-Flight Tandem Mass Spectrometry

Abstract

) has been used for the identification and structural determination of steroids. Steroidal glycoalkaloids extracted from natural plants are identified using electrospray ionization (ESI) in ion trap (IT) MS,1) and steroid glucuronides are analyzed using a triplequadrupole (Q) instrument with a turbo ion-spray source. 2)An acetylated cholestane diglycoside, OSW-1 (1), (3b,16b,17a-trihydroxycholest-5-en-22- one 16-O-{O-[2-O-(4-methoxybenzoyl)-b -D-xylopyranosyl]-(1→3)-2-O-acetyl-a-L-arabinopyranoside}), was isolated by Sashida and co-workers as the main constituent from the bulbs of Ornithogalum saundersiae (Liliaceae) and found to show potent cytotoxicity against a variety of malignant human tumor cells.3) OSW-1 exhibits low toxicity against normal cells but inhibits the growth of malignant cells and is 10-100-fold more potent than clinically established anticancer agents such as mitomycin C, doxorubicin, camptothecin, and paclitaxel.4) Fuchs and colleagues proposed that the active intermediate might be a 22-oxocarbenium ion, which could be generated from 22-carbonyl and 16a-hydroxy moieties.5) Yu and colleagues have recently reported that both 22-methylene and 23-heteroatom (O, S, NH) analogues of OSW-1 were as potent as the parent natural products against the growth of tumor cells. 6) However, the precise mechanism by which OSW-1 exerts its effects remains unclear. A further study of the structure-activity relationships of OSW-1 analogues having heterocyclic rings, such as thiophene and thiazole, at the side chain was therefore designed. We have succeeded in the syntheses of OSW-1 (1) and one of its analogues, ThOSW-1, (20S,23Z,25Z)-3b ,16b ,17a -trihydroxy-23,26-epithiocholesta-5,23,25-trien-22-one 16In a previous study, we investigated the optimum LC/MS conditions to obtain useful ions required to identify structures using ESI and atmospheric-pressure chemicalionization (APCI)-QMS. 9)Cardiotonic steroids are classified into two groups based on the type of unsaturated lactone ring substituted at the 17b-position of the steroidal skeleton, i.e., cardenolides with a five-membered lactone ring and bufadienolides with a sixmembered lactone ring, respectively. Toad steroids isolated from Bufo sp. have a six-membered lactone ring and are divided into two groups according to the form at the 3b-position of the steroid: bufogenin as an unconjugated steroid; and bufotoxin as a steroid conjugate. From ancient times, the skin secretions of native toads, known as ch'an su in China and as senso in Japan, have been employed as cardiac or diuretic agents, and the main cardiotonic constituent in toad venom has been elucidated to be bufotoxin. 10) Marinobufotoxin (3) with a suberoylarginine ester at the 3b-position is one of the main cardiotonic constituents. In addition to this, homologues with other dicarboxylic acids, i.e., pimeric acid, succinic acid, or adipic acid, and analogues with other amino acids, i.e., histidine or glutamine, are known to occur. In addition, other conjugate forms, i.e., sulfates or glucuronides, are...

Production of free methylarginines via the proteasome and autophagy pathways in cultured cells

“…The structural identities of the HPLC-fractionated peaks corresponding to ADMA and SDMA were confirmed by MS, MS/MS, and MS/MS/MS sequential analyses with MALDI-QIT-TOF/ MS (Fig. 1B) (19). The present methods resulted in fairly good recovery rates (>80%), and linear regression analysis showed significant linearity of the calibration curve (Fig.…”

“…External calibrations were achieved using the standard reagents Ang II [monoisotopic mass (M+H) + = 1046.5], and bradykinin 1-7 [(M+H) + = 757.4]. Tandem mass experiments with MALDI-QIT-TOF/MS n (n=1, 2, or 3) were performed using argon gas as the collision gas on an AXIMA-QIT mass spectrometer (Shimadzu, Kyoto, Japan) equipped with a nitrogen laser (337 nm) with the collision-induced dissociation control value set at 160 (19).…”

Structural features of polyacylated anthocyanins using matrix‐assisted laser desorption/ionization and electrospray ionization time‐of‐flight mass spectrometry

“…[13] In MALDI-MS analysis of steroid glycosides, the affinity of sodium cations for glycosides was greater than that of protons. [18] In addition, the fragment ions at m/z 995, 833, and 671, derived from subsequent losses of 162 u from the ion at m/z 1157, appeared to correspond to the elimination of glucose and caffeic acid, indicating that further fragmentation of the caffeoylsophoroside substituent occurred in MALDI-MS. Because the mass accuracy was only within 0.4 u, caused by irregular flight distance probably originating from slight arching of the sample plate or a slight variation in the analyte thickness, it was not possible to differentiate glucose (C 6 H 10 O 5 , monoisotopic mass 162.0528) and caffeic acid (C 9 H 6 O 3 , monoisotopic mass 162.0317).…”

“…In our previous studies on the structural elucidation of biologically active steroid glycosides and steroid conjugations, ESI-MS and MALDI-MS techniques were confirmed to be useful. [17,18] In determining the structures of newly isolated acylated anthocyanins, MS plays an essential role. A sector mass spectrometer equipped with a fast atom bombardment (FAB) ionization apparatus was used for the identification of the molecular ions and determination of the elemental compositions of pigments 1-5 extracted by the authors from Ipomoea purpurea, Petunia guarapuavensis, Senecio cruentus, and Bletilla striata.…”

Proteomics of the Rodent Malaria Parasite Using Matrix-Assisted Laser Desorption/Ionization Quadrupole Ion Trap Time-of-Flight Tandem Mass Spectrometry

“…In the present study, the application of MALDI-quadrupole ion trap (QIT)-TOF tandem MS, which is used for structural determination of conjugated compounds 9 or posttranslational modifications, 10 for the analysis of native, not recombinant, proteins from P. berghei strain NK 65 parasite was demonstrated. Protein identification using both the peptide mass fingerprint (PMF) and continuous MS/MS ion search methods via the MSDB20060831 and NCBI20111206 (except 20111203 for spot No.…”