Phycobiliproteins represent a considerable fraction (upto 50%) of the total cellular proteins of cyanobacteria (Bennett and Bogorad, 1973; Tandeau de Marsac, 1977). Besides their crucial function of harnessing solar energy for algal food production, phycobiliproteins have immense commercial value as natural pigments for beverages, dry beverage mixtures and natural dyes (Takano et al., 1992). The use of phycobiliproteins in fluorescence microscopy, fluorescence immunoassay and as phycoflours has been well realized (Richmenn, 1990; Rodriquez et al., 1989). Further, phycocyanin, the major phycobiliproteins has been shown to possess anticancer activity to stimulate immune system and to treat ulcers and haemorrhoidal bleeding (Rodriquez et al., 1989). Phycocyanin is commercially produced in Japan from Spirulina platensis and used for coloring foodstuffs and also in cosmetics such as eye shadow, eyeliner and lipstick preparations (Venkataraman, 1989).

Analysis of phycobilins in four species of cyanobacteria reveals that in cyanobacteria the concentration of phycobilins is species-dependent. Among the four cyanobacteria tested with in the present study a gradation of phycobilin concentration was found to exist (Table 1). Oscillatoria had the lowest concentration of pigments, while Scytonema showed a six-fold higher content compared to the former. Based on the variations observed in the phycobilin concentrations, it was possible to categorize the four cyanobacterial species in the order of their decreasing phycobilin content as Scytonema > Nostoc > Westiellopsis > Oscillatoria.

The concentration and composition of phycobiliproteins have been reported to vary depending on the environmental conditions like nutrients, temperature, light quality and quantity (Halldal, 1970; Miller and Holt, 1977). Therefore, it was thought to be worthwhile to employ stress factors as tools in order to enhance the phycobilin yield potential of cyanobacteria. A physical stress namely high light condition (500 µmol m^-2 s^-1 PPFD) and a chemical stress (herbicide glyphosate at 40 and 80 µM) where chosen. Results presented in Table 2 show that except in Oscillatoria, in the remaining three organisms namely Scytonema, Nostoc and Westiellopsis, high light conditions have a promotory effect on the synthesis of phycobilins. Particularly, in Westiellopsis the high light mediated synthesis of phycobilins was 2 to 5 fold higher compared to the control. Among the four phycobilins analyzed phycocyanin and allophycocyanin registered profound increase compared to phycoerythrin and phycobiliproteins (Table 2). Bryant and Cohen-Bazire (1981) and Bryant (1982) have noted that more than one set of phycocyanin subunits was synthesized in light adapting cyanobacteria and such findings implied that multiple phycocyanin genes are differentially controlled under different light conditions. Our results also substantiate this proposal (Table 2). Further, the reduction in the phycobilins noticed in Oscillatoria proves that this organism is highly sensitive to high light conditions.

The chemical stress glyphosate treatment has brought in a very high level of phycobilin synthesis in Oscillatoria only at the lower do employed (40 µM). Nevertheless, in the remaining three cyanobacteria it does not significantly alter the pigment concentration (Table 2). Further, on the contrary, the higher dose of glyphosate treatment (80 µM), has enhanced the phycobilin concentrations in Scytonema, Nostoc and Westiellopsis. But in Oscillatoria this dose has reduced the magnitude of enhancement in the phycobilin contents compared to the lower dose treatment (Table 2). An enhanced level of phycobilisome pigments under glyphosate treatment has been reported from our previous experiments (Ravi and Balakumar, 1998; Balakumar et al., 2000). Cyanobacteria showing enhanced levels of pigments under glyphosate treatment on the whole, express a higher level of resistance to glyphosate due to the possession of one or more kinds of adaptive mechanisms. The enhanced level of phycobilins in Scytonema, Nostoc and Westiellopsis under the higher dose of glyphosate treatment indicates that these organisms also can be regarded as glyphosate resistant ones. However, this proposal remains to be validated further. The differential responses of Oscillatoria to varying levels of glyphosate stands unclear at this point which call for further experimentation.

Simultaneous exposure of plants to multiple stresses in many cases has been proved to be more beneficial to plants compared to either of the stresses given alone by bringing in better effects (Balakumar et al., 1993). Results obtained in this present study concord with this hypothesis. A combination treatment of high light along with glyphosate has evoked a promotory effect in the synthesis of phycobilins in all the four species of cyanobacteria investigated (Table 3). Under this treatment also Westiellopsis and Scytonema have registered their higher level of response than Nostoc and Oscillatoria. However, the combination of stresses brought in only a lesser extent of enhancement in pigment concentration compared to high light given alone. On the other hand, Nostoc showed a strikingly different response under the combination of stresses by an enhanced level of pigments, while high light alone had an inhibitory effect on pigment synthesis (Table 2 and 3). Therefore, the combination of two stresses could be effective and useful in scaling up the phycobiliprotein potential at least in Oscillatoria, while exposure to high light may be a suitable technology for Scytonema, Nostoc and Westiellopsis. The reduction in phycobilins in these three organisms under combined stresses compared to high light stress given alone indicates that the action of light in the synthesis of phycobilins is hindered by glyphosate. The mechanism of such an interaction is not clear at this point, which remains to be elucidated further.

Acknowledgements

The authors thank the Research and Development Committee of the American College, Madurai 625 002, India, for financial support.