The hydrogen bonding of cytosine with guanine: calorimetric and 1H-nmr analysis of the molecular interactions of nucleic acid bases

The enthalpy of hydrogen-bond formation between guanine (G) and cytosine (C) in o-dichlorobenzene and in chloroform at 25°C has been determined by direct calorimetric measurement. We derivatized 2′-deoxyguanosine and 2′-deoxycytidine at the 5′- and 3′-hydroxyls with triisopropylsilyl groups; these groups increase the solubility of the nucleic acid bases in nonaqueous solvents. Such derivatization also prevents the ribose hydroxyls from forming hydrogen bonds. Consequently, hydrogen-bond formation in our system is primarily between the bases, and to a lesser extent, between base and solvent, and can be measured directly with calorimetry. To obtain the data on base-pair formation, we first took into account the contributions from self-association of each base, and where possible, have determined the ΔH of self-association. From isoperibolic titration calorimetry, our measured ΔH of C₂ formation in chloroform is ?1.7 kcal/mol of C. Our measured ΔH of C:G base-pair formation in o-dichlorobenzene is ?6.65 ± 0.32 kcal/mol. Since o-dichlorobenzene does not form hydrogen bonds, the ΔH of C:G base-pair formation in this solvent represents the ΔH of the hydrogen-bonding interaction of C with G in a nonassociating solvent. In contrast, our measured ΔH of C:G base-pair formation in chloroform is ?5.77 ± 0.20 kcal/mol; thus, the absolute value of the enthalpy of hydrogen bonding in the C:G base pair is greater in o-dichlorobenzene than in chloroform. Since chloroform is a solvent known to form hydrogen bonds, the decrease in enthalpic contribution to C:G base pairing in chloroform is due to the formation of hydrogen bonds between the bases and the solvent. The ΔH of hydrogen bonding of G with C reported here differs from previous indirect estimates: Our measurements indicate the ΔH is 50% less in magnitude than the ΔH based on spectroscopic measurements of the extent of interaction. We have also observed that the enthalpy of hydrogen bonding of C with G in chloroform is greater when G is in excess than when C is in excess. This increased heat is due to the formation of C:G_{n > 1} complexes that we have observed using ¹H-nmr. Although C:G₂ structures have previously been observed in triple-stranded polymeric nucleic acids, higher order structures have not been observed between C and G monomers in nonaqueous solvents until now. By using monomers as a model system to investigate hydrogen-bonding interactions in DNA and RNA, we have obtained the following results: A direct measurement of the ΔH of hydrogen bonding in the C:G complex in two nonaqueous solvents, and the first observation of C:G_{n > 1} complexes between monomers. These results reinforce the importance of hydrogen bonding in the stabilization of various nucleic acid secondary and tertiary structures.     

Volubilin,a new isoflavone-C-glycoside from Dalbergia volubilis flowers

Dalbergia volubilis flowers after successive extraction with light petroleum, benzene and alcohol yielded β-sitosterol, stigmasterol, biochanin-A and a new isoflavone-C-glycoside named volubilin. The glvcoside on boiling with HI and phenol gave genistein and on ferric chloride oxidation rhamnose. The compound was identified as 8-C-rhamnopyranosyl-genistein-4′,7-dimethyl ether by means of chemical reactions as well as from spectral data.