The objective of this study was to clarify the factors that influence saponin biosynthesis, with an emphasis on protodioscin, a major saponin compound in white asparagus spears. We investigated the effects of methyl jasmonate (MeJM) treatment on protodioscin content in white spears harvested by spring semi-forced and winter-forced cultivation. Protodioscin content was determined using high performance liquid chromatography equipped with an evaporative light scattering detector. Protodioscin was mainly localized in the lower region of the spears. In the spring semi-forced cultivation system using the film-cover method, the protodioscin content in white spears harvested from the MeJM plot was significantly higher than in those harvested from the control plot throughout the sampling period. MeJM also significantly enhanced protodioscin biosynthesis in a dose-dependent manner in the experiment conducted under winter-forced cultivation. These results suggest that saponin biosynthesis in white spears is associated with plant defensive reactions triggered by jasmonate. In addition, bitterness in film-covered white spears could be artificially controlled in commercial fields by MeJM treatment.