111 SINGLE EQUINE EMBRYO LIPID FINGERPRINTING BY MASS SPECTROMETRY
R. F. Gonçalves A , C. R. Ferreira B , C. M. B. Orlandi C , V. C. Sartori C , H. N. Ferreira C , F. C. Gozzo D , S. A. Saraiva D , E. J. Pilau D and M. N. Eberlin BA Department of Animal Reproduction, College of Veterinary Medicine and Animal Science, São Paulo, SP, Brazil;
B ThoMSon Mass Spectrometry Laboratory, Institute of Chemistry, University of Campinas, Campinas, SP, Brazil;
C Veterinary Medicine College, Faculdade Integrado de Campo Mourão, Campo Mourão, PR, Brazil;
D Dalton ThoMSon Mass Spectrometry Laboratory, Institute of Chemistry, University of Campinas, Campinas, SP, Brazil
Reproduction, Fertility and Development 23(1) 160-161 https://doi.org/10.1071/RDv23n1Ab111
Published: 7 December 2010
Abstract
Matrix-assisted laser desorption-ionization mass spectrometry (MALDI-MS) allows the lipid profile study of individual mammal embryos. The data collection is rapid, highly sensitive, can tolerate some level of impurities, and is easy to interpret. The aim of this study was to report the lipid profile obtained from a single equine embryo by MALDI-MS. Follicles ≥30 mm in diameter were monitored daily until ovulation (Day 0). The insemination was performed close to ovulation with fresh diluted semen, and the embryo recovery was performed on Day 9 (D9) post ovulation. The equine embryo was placed in 50/50 (v/v) methanol/phosphate buffer solution and transported at 4°C to the laboratory. MALDI-MS spectra were acquired in the positive ion mode using MALDI Synapt HDMS mass spectrometer (Waters, Manchester, UK) m/z 700–950 range. The sample was coated with a 1.2 μL matrix of 2,5-dihydroxybenzoic acid (DHB) 1.0 mL L–1 in methanol. Due to the equine embryo volume, it was possible to divide it and get two mass spectra, which were identical. Spectra processing was performed using the MassLynx 4.0 software (Waters, Manchester, UK). It was observed the presence protonated and sodiated species of sphingomyelins (SM), phosphatidylcholines (PC), and triacylglycerols (TAG). The most intense ions assigned by comparison with data obtained from bovine embryos were m/z 723.5 [PC (34:1) and loss of N(CH3)3]+, 725.5. [SM (16:0) + Na]+, 754.6 [PC (32:1) + Na]+, 778.6 [PC (36:1) + Na]+, 780.6 [PC (34:2) + Na]+ or [PC (36:5) + H]+, 782.6 [PC (36:4) + H]+ or [PC (34:1) + Na]+, 788.6 [PC (36:1) + H]+, 806.6 [PC (38:6) + H]+ or [PC (36:3) + Na]+, 808.6 [PC (38:5) + H]+ or [PC (36:2) + Na]+, 810.6 [PC (38:4) + H]+ or [PC (36:1) + Na]+, 907.7 [TAG (54:3) + Na]+ and 909.7 [TAG (54:2) + Na] +. Regarding the lipid profile by MALDI-TOF previously reported for oocytes and embryos of several species (Ferreira et al. 2010, J. Lipid Res., 51, 1218–1227), it detected similar lipid species, but with different relative intensities. Because of the single equine embryo volume and MALDI-MS technique sensitivity, we intend to observe if there will be differences between the lipid profile of the inner cell mass and trophoblast in the future. The analysis of a greater number of embryos as well as different development periods and MS/MS experiments will contribute to building a database of lipid profiles that allows a better understanding of the lipid profile physiology in equine embryos and the meaning of differences among other mammalian embryos.
FAPESP (São Paulo Research Foundation).
