Studies On The Spectral Analysis Methods Of Mongolian Piperlonguminine And Its Interction With Bovine Serum Albumin

Piperlonguminine(GBN) is the Piperine derivatives extracted from the immature ear of Long pepper by extraction and separation, which can fall hematic fat obviously So far there is no effective method for determinating the GBN in biological samples. Therefore it is very necessary to establish a rapid, simple and accurate analysis method for the determination of GBN in biological samples. In this research work taking GBN as the research object, new methods of GBN detection in biological samples were established and the interactions between GBN and bovine serum albumin (BSA) was studied by fluorescence spectrometry and ultraviolet absorption spectrometry. It has important theoretical significance for understanding the metabolic processes of GBN in biological body and illuminating its pharmacological effects in the body.In this researche, firstly taking rare earth complexes ArsenazoⅢ-La(Ⅲ) as spectral probe and basing on the complexes formed by ArsenazoⅢ-La(Ⅲ) and GBN which composition ratio was1:1:2under the acidic condition, the Ultraviolet-visible Absorption spectrum and Resonance Rayleigh Scattering spectrum of ArsenazoⅢ-La(Ⅲ) and GBN was studied. The Fading Spectrophotometric Method and Resonance Rayleigh Scattering Method was established to determinate the GBN in biological samples respectively. The range of linearity was0.02733～0.5466μg/mL and0.02733～5.466μg/mL and the detection limit was0.02206μg/mL and0.00859μg/mL respectively. To determinate GBN in biological samples by this system, it was increased in sensitivity and interference immunity which compared with pre-existing methods for GBN detection. Then according to the fluorescence characteristics of GBN and its fluorescence intensity could be sensibilized by surfactant tween-20, the micellar sensitization fluorescence method was set up to determinate GBN. The range of linearity was0.02733～8.199μg/mL and the detection limit was0.001690μg/mL. It could be used to determinate GBN in biological samples by appropriate separation. On this basis, using synchronous fluorescence technology to eliminate the interference of biological samples’background fluorescence on GBN’s fluorescence spectrum, a new method was established for detecting GBN in serum directly. The range of linearity was0.005466～3.2796μg/mL and the detection limit was0.001197μg/mL. The fluorescence determination of GBN in rats serum was direct and quick. The proposed fluorescence method had high sensitivity and good selectivity and the actual measurement result was satisfactory.In this work, the interaction between Piperine(PP), Curcumin (Cur) and GBN was also studied and its influence on GBN and BSA was discussed respectively under the simulated physiological conditions by ultraviolet-visible absorption spectrometry and fluorescence spectrometry. It turned out that the interaction processes of GBN, PP, Cur with BSA were static quenching and the association constants were Ka(BSA-PP)> Ka(BSA-GBN) and Ka(BSA-GBN)>Ka(BSA-Cur). According to the combination parameters, the operating distance and corresponding thermodynamic parameters under different temperatures, it made sure that the reaction type were electrostatic force, hydrophobic force and hydrophobic force respectively. And the binding sites of BSA were site Ⅰ, site Ⅱ, site Ⅱ respectively by the marker competition experiments. The BSA conformation was affectet by GBN, PP, Cur which determined by ultraviolet absorption spectrum and synchronous fluorescence spectrum and the binding sites were more closer to tryptophan residues. The collaborative work among multivariate ligands showed that the combination between GBN, PP, Cur alone with BSA was negative collaborative. When PP, Cur was coexisted with GBN, it was without collaborative to the binding between GBN and BSA. But the increasing of association constant K(BSA-PP-GBN)>K(BSA-GBN)、K(BSA-Cur-GBN)> K(BSA-GBN) revealed that the effects of PP and Cur on the binding of GBN and BSA were addition. The Stokes shifts and fluorescence intensity differences of three-dimensional fluorescence spectrometry explained the interactions between GBN and PP, Cur further.