Induction of ovulation and superovulation in mares using equine LH and FSH separated by hydrophobic interaction chromatography

Abstract

Pharmacological control of reproduction in mares requires the use of equine gonadotrophins to avoid induced immunological resistance. Crude equine gonadotrophins (CEG) have been used but the presence of equine luteinizing hormone (eLH) and follicle-stimulating hormone (eFSH) in CEG has led to disappointing results in superovulation studies. Separation of eLH and eFSH activities from CEG is necessary to overcome this problem. The hydrophobic properties of the two hormones were sufficiently different to permit their separation by hydrophobic interaction chromatography (HIC) on a phenyl Sepharose matrix. Good yields of separate FSH and LH fractions were readily obtained by stepwise elution and the method was adapted for large scale preparations of enriched fractions of eLH and eFSH. Two experiments were performed in vivo to evaluate the biological activity of the HIC fractions. Experiment 1 showed that biological activity of the LH fraction in inducing ovulation of preovulatory follicles was similar to that obtained with CEG, indicating that LH bioactivity was not altered by HIC. Experiment 2 demonstrated that biological activity of the FSH fraction was identical (as far as rate of ovulation was concerned) to that of CEG in superovulating mares, indicating that FSH activity was also not altered by HIC. Although we have not obtained better results with the separate equine gonadotrophins than with CEG, it is potentially advantageous to use preparations with single activity to obtain a controlled balance of FSH and LH activity. The HIC technique was chosen because it could easily be scaled up to provide the large amounts of the separate hormones needed for the treatment of a large number of mares.