Pyrazine derivatives in cigarette smoke inhibit hamster oviductal functioning

BACKGROUND Our past studies have shown that cigarette smoke inhibits oviductal functioning in vivo and in vitro. The goals in this study were to identify pyrazine derivatives in cigarette smoke solutions that inhibit ciliary beat frequency, oocyte pickup rate, and infundibular smooth muscle contraction in the hamster oviduct and to determine their lowest observable adverse effect levels (LOAELs) using in vitro bioassays. METHODS MS smoke solutions were fractionated using solid phase extraction cartridges and the fractions were both tested on the hamster oviduct in vitro and analyzed by gas chromatography-mass spectrometry to identify individual pyrazine derivatives. Commercial pyrazine standards were purchased, assayed for purity, and tested in dose-response studies on hamster oviducts. The LOAEL and efficacy were determined for each compound in the in vitro bioassays. Statistical significance was determined using the Student's t-Test where p < 0.05. RESULTS The LOAELs for the most inhibitory pyrazine derivatives in the ciliary beat frequency, oocyte pickup rate, and infundibular smooth muscle contraction assays were as follows: for pyrazine (1 picomolar, 10 picomolar, and 1 nanomolar); for 2-methylpyrazine (1 picomolar, 10 picomolar, and 10 picomolar); and for 2-ethylpyrazine (1 picomolar, 10 picomolar, and 1 picomolar). Six of the seven pyrazine derivatives tested (pyrazine, 2-methylpyrazine, 2-ethylpyrazine, 2-methoxy-3-methylpyrazine, 2,5-dimethylpyrazine, and 2,3,5-trimethylpyrazine) were inhibitory in picomolar or nanomolar doses in all three bioassays, while the seventh derivative, 2,6-dimethylpyrazine, had LOAELs in the nanomolar to micromolar range. CONCLUSION This work shows that very low doses of pyrazines significantly inhibit proper oviductal functioning, raising questions regarding the safety of these compounds in cigarettes and other consumer products.