Phenologic variation of major triterpenoids in regular and white Antrodia cinnamomea

In this study, we analyzed the chemical profiles of ethyl acetate extracts from a regular form, a natural occurring white variant, and a whitened variant induced by blue light at different growth stages. The TLC and HPLC analyses indicated that chemical profiles of ethyl acetate extracts from the regular form were getting more complex as the fungus grew while the white variants remained less complex (Fig. 3). This patterns became obvious when the regular form were 6 weeks old when the process of fructification started. This phenologic changes of the chemical profiles coincided with the morphological changes of the regular form from mycelium to fruiting body (Chu et al. 2010). There were no significant morphological changes in the naturally occurring white variant and the whitened variant. It indicates that their chemical profiles changed dramatically during fungal fructification and may produce novel medicinal effects.

A total of 39 triterpenoid compounds had been identified and structurally elucidated from A. cinnamomea based on the review of Geethangili and Tzeng (2011). These triterpenoid structures have an ergostane skeleton (Antcin A, C, K, Zhankuic acid A, B, and C in this study) or a lanostane skeleton (Sulphurenic acid, Dehydrosulphurenic acid, Eburicoic acid, and Dehydroeburicoic acid in this study) The ergostane-type triterpenoids are mostly found in fruiting bodies and rarely reported in mycelia when the triterpenoids of a wild A. cinnamomea fruiting body and those of a submerged culture were compared (Geethangili and Tzeng 2011). It is generally believed that ergostane-type triterpenoids are produced in fruiting bodies and lanostane-type triterpenoids exist both in fruiting bodies and in mycelia. However, this acknowledge of triterpenoid distribution in A. cinnamomea may not be true when A. cinnamomea was cultured in artificial agar-plate media. In our study, both ergostane-type and lanostane-type triterpenoids were detected in young mycelia and old mycelia with emerging of fruiting body. Most of the time, production of the ergostane-type triterpenoids was dramatically increased when the cultures were at 4–6 weeks old, but lanostane-type triterpenoids did not show a clear trend in quantity changes at this point (Tables 1, 2, 3). This indicates the biosynthesis of ergostane-type triterpenoids may be accelerated during A. cinnamomea fructification, but not lanostane-type triterpenoids. The lanostane-type triterpenoids may be involved in house-keeping during the growth and development of A. cinnamomea.

Differentiation and secondary metabolism are correlated processes in fungi that respond to light (Bayram et al. 2008). We analyzed ten key triterpenoids for their relative quantities during eight weeks of growth. In general, both naturally occurring white variant and induced whitened strains shared a higher pattern similarity than either with the regular strain. However, the blue-light induced whitened strain did contain a strong characteristic peak of antcin K, which was not shown much in the naturally occurring white strain (Fig. 5). This indicates the metabolism of triterpenoids in both the naturally occurring variant and the whitened strain may not carry the same pathways despite of both having white colonies. Antcin K which is considered a characteristic compound in A. cinnamomea fruiting bodies may be an early fructification-specific triterpenoid. Further fructification, which occurred in the regular strain, was seemingly blocked by blue-light treatment.

The effects of light treatment on fungal growth of various species had been studied earlier but with conflicted results. Chen and Dickman (2002) had shown that light treatment may induce a Colletotrichum trifolii TB3 kinase gene expression and help hyphal elongation and branching. However, the treatment of blue light radiation inhibited the apical growth of Tuber borchii mycelium by induction of a photoreceptor, Tbwc-1, gene expression (Ambra et al. 2004). Interestingly, in current studies, we found both phenomena occurred in our observation on growth of A. cinnamomea. The treatment of blue light radiation on regular A. cinnamomea increased the growth of fungal mycelium during the first ten days of inoculation, and then inhibited its growth thereafter (Fig. 2). This peculiar phenomenon indicates a more complex physiological interaction between fungal growth and light radiation. This phenomenon also suggests to us an improved procedure of culturing A. cinnamomea by treating cultures with light radiation for ten days before placing them in the dark.