The screened compounds from Ligustri Lucidi Fructus using the immobilized calcium sensing receptor column exhibit osteogenic activity in vitro.

Calcium sensing receptor (CaSR) has become the novel target of treating osteoporosis with herbal medicine Ligustri Lucidi Fructus (LLF), however, the bioactive compounds responsible for anti-osteoporosis are hard to clarify due to the complexity and diversity of chemical constituents in it. Herein, the immobilized CaSR column was packed with stationary phase materials, which were derived from integrating CLIP-tagged CaSR directly out of crude cell lysates onto the surface of silica gels (5.83 mg/g) in a site-specific covalent manner. The column had a great specificity of recognizing agonists and kept a good stability for at least 3 weeks. The two compounds from LLF extract were screened and identified as olenuezhenoside and ligustroflavone using the immobilized CaSR column in conjunction with mass spectrometry. Molecular docking predicted that both compounds were bound in venus flytrap (VFT) domain of CaSR by the formation of hydrogen bonds. Cellular results showed that both compounds exhibited the distinct osteogenic activity by enhancing the proliferation, differentiation and mineralization of osteoblastic cells. Our study demonstrated that, the immobilized protein column enables to screen the bioactive compounds rapidly from herbal extract, and the newly discovered natural product ligands towards CaSR, including olenuezhenoside and ligustroflavone, will be the candidates for the treatment of osteoporosis.

Solid-phase extraction with the functionalization of calcium-sensing receptors onto magnetic microspheres as an affinity probe can capture ligands selectively from herbal extract.

Magnetic solid phase extraction with the functionalization of protein onto micro- or nano-particles as a probe is favorable for the discovery of new drugs from complicated natural products. Herein, we aimed to develop a rapid method by immobilizing halogenated alkane dehalogenase (Halo)-tagged calcium-sensing receptor (CaSR) directly out of crude cell lysates onto the surface of magnetic microspheres (MM) with no need to purify protein. Thereby we achieved CaSR-functionalized MM for revealing adsorption characteristics of agonist neomycin and screening ligands from herbal medicine Radix Astragali (RA). About 43.87 mg CaSR could be immobilized per 1 g MM within 30 min, and the acquired CaSR-functionalized MM showed good stability and activity for 4 weeks. The maximum adsorption capacity of neomycin on CaSR-functionalized MM was determined as 4.70 × 10-4 ~ 3.96 × 10-4 mol/g within 277 ~ 310 K, and its adsorption isotherm characteristics described best by the Temkin model were further validated using isothermal titration calorimetry. It was inferred that CaSR's affinity for neomycin was driven by electrostatic forces in a spontaneous process when the system reached an equilibrium state. Moreover, the ligands from the RA extract were screened, three of which were assigned as astragaloside IV, ononin, and calycosin based on HPLC-MS. Our findings demonstrated that the functionalization of a receptor onto magnetic materials designed as an affinity probe has the capability to recognize its agonist and capture the ligands selectively from complex matrices like herbs.

Macromolecule-to-amphiphile conversion process of a polyoxometalate-polymer hybrid and assembled hybrid vesicles.

We report our findings on the macromolecule-to-amphiphile conversion process of a polyoxometalate-polymer hybrid and the assembled hybrid vesicles formed by aggregation of the hybrid amphiphile. The polyoxometalate-polymer hybrid is composed of a polyoxometalate (POM) cluster, which is covered by five tetrabutylammonium (Bu(4)N(+)) countercations, and a polystyrene (PS) chain. Through a cation-exchange process the Bu(4)N(+) countercations can be replaced by protons to form a hybrid amphiphile composed of a hydrophilic, protonated POM cluster and a hydrophobic PS chain. By implementing a directed one-dimensional diffusion and analyzing the diffusion data, we confirmed that the diffusion of solvated protons rather than macromolecules or aggregates is the key factor controlling the conversion process. Once the giant hybrid amphiphiles were formed, they immediately assembled into kinetically favored vesicular aggregates. During subsequent annealing these vesicular aggregates were transformed into thermodynamically stable vesicular aggregates with a perfect vesicle structure. The success in the preparation of the POM-containing hybrid vesicles provides us with an opportunity of preparing POM-functionalized vesicles.

Ribbonlike assembly of molecules composed of fulleropyrrolidine and PUA dendron.

The hybrid molecules composed of fulleropyrrolidine and poly(urethane amide) dendron of the third generation have been synthesized, and their self-assembling in THF/water mixtures has been studied. By TEM, AFM, and SAXS methods it was established that molecules form ribbonlike aggregates with the ordered layers of fullerene derivatives situated inside of the ribbons. Microscale length and high ratio width/thickness were detected for the ribbons. The ability of amides and urethanes to initiate a coupling of dendron-fragments by means of hydrogen bonding has been considered as mainly responsible for anisotropic interaction of hybrid molecules and for the ribbonlike aggregates formation in THF/water mixtures.