**2.7 Transoformation of soybean oil, sunflower oil to bioadhesive**

290 Soybean – Genetics and Novel Techniques for Yield Enhancement

lower concentrations displayed only the micellar structures. In the first stage, we visualized the transformation of soybean oil to micelle formation similar to the micelle of experimental flasks receiving 2.5 and 5.0% concentrations. Initial micellar formation could be due to agitation and the mineral salts present in the medium. Increasing the incubation period, further (after 6 days) results with vesicle formation and then to gellation. The formation of vesicle and gellation could be due to the primary aggregation of micelles. Similar to the report on self-assembly of peptides, in the present study, presence of glucose, peptone and the prevailing temperature (37C) may initiate the primary aggregations. In addition, lecithin concentration and the phosphate moiety induce gellation and stabilization of

When the incubation period further increased to beyond 10 days we observed a formation of a single macro-sized organogel. The gel was spherical in shape with 2 cm in diameter. The inner structure of the cross sections showed multiple layers arranged like onion rings with no hallow space with tissue like appearance and feel in the center core. The thickness of the outer membrane was measured as 1 mm. According to Ai *et al*., (2003) and Liu *et al*., (2003) the layered assembly has molecular scale thickness and a few tens of layers can be easily achieved and the total layered area can be extended above the micrometer scale. Similarly, we observed continuous layering results with a macro-size organogel of 2 cm in diameter. Since, the directional and differential orientation of self-assembly decides the final size and structure of the secondary aggregates, in the present study, layering continues with directional orientation resulting with spherical size. Additional hydrogen bonding and the external agitation force may be responsible for the directionality, and the weak intermolecular forces (Van der Waals attractions) play the major role in the force balance, which results with multiple layering of bilayers in the secondary aggregations. Other than the above said driving forces, pH of the medium (6.0 0.5), presence of divalent cations and anions (transition metal salts), and temperature may also instigate the secondary aggregations and the co-operation between the primary and the secondary aggregations. There is a possibility of two different modes of primary aggregations for the existence of macrosized organogel. If the primary aggregation was a layered type, then the secondary aggregation could be a multi-layered with the space in between them able to hold either water or lipophilic compound depends on the nature of gel. In other words, if the selfaggregation was a fusion of micelles then the flower like aggregations takes place results

with the circular/ spherical layered structures with same kind of holding nature.

Fig. 6. Macrosized self-assembled organogel from soybean oil.

micelles.

In the present study, dimerization of fatty acids of triglycerides have been realized during microbial growth. Growing the chosen marine microorganism in the medium increases the multiplication of cells and the doubling time (30 minutes). Though, triglycerides were given at varied concentrations (2 to 10 weight percent), only with >5.0 weight percent concentration, visible changes in the oil phase of the medium were observed.

At >5.0 percent concentration of oil we visualized the transformation of oil phase within 24 to 48 h of incubation. This might by due to the rapid *in situ* hydrolysis of triglycerides by the lipolytic enzymes produced by the organisms. Similarly, an increase in glycerol content in the cell free broth also substantiates the enzymatic process of the cleavage of triglycerides and the presence of a biosurfactant with appreciable surface activity of cell free broth may also contribute to the formation of thread like structures.

The presence of free fatty acids at trace levels evidenced throughout the experimental period in the form of oleic or linoleic acid further confirmed the hydrolysis of oil and the percentage of linoleic acid has been found at higher level compared to oleic acid. The formation of thread like structures further condensed to develop in to a solid mass with adhesive nature when the incubation period increases beyond 120-h.

*In situ* generation of biosurfactants plays a vital role in the condensation and polymerization as reported by Kestelman and Veselovsky (2001). The increased adhesive nature observed in the product of the present study might be due to the biosurfactants available in the broth during the formation of the adhesive product and its involvement in the intramolecular network of the product. Markevich et al., (1986) detailed the role of surfactant in increasing the adhesive nature of the product, where, they employed synthetic surfactants.

The thread like structures formed during the growth of the organism has been considered as the dimers of fatty acids or triglycerides polymers and these dimers are the precursors for the product formation, however, we couldn't ascertain the nature of dimers formed. Followed by dimerization, the reaction may further preceded and provide the product with adhesiveness. Dimerization followed by product formation might be affected by the presence of (i) biosurfactant; (ii) available amino acids (iii) unspent metal ions (iv) free glycerol and (v) monomers of triacylglycerols in the medium. With regard to dimerization in aqueous medium, Lyons (1969), Wheeler and Godfrey (1974) and Isbell (1994) reported less percentage of water increases the yield of dimers. However, in the present study, dimerization was evidenced in the aqueous medium. The following figure illustrate the adhesive nature of the supramolecular self-assembly structures of soybean (A) and sunflower (B).

Fig. 7. Bioadhesive from soybean oil and sunflower oil.

Transformation of Soybean Oil to Various Self-Assembled Supramolecular Structures 293

Fig. 9. Schematic representation of different self-assembled supramoelcular structures using

plant triglycerides with microbes, with microbial products and by simple physical

treatments.
