We have synthesized nanoporous silica, SBA-15, in the 1 μm size range with the pore diameter of 7.6 nm. The redox enzyme horseradish peroxidase (HRP) was entrapped in the pores to form nanostructured hybrid materials. The catalytic activity of free and immobilized enzyme was first compared at room temperature. Details of the enzyme kinetics including the apparent Michaelis constant (K M) and maximum rate (V max) were determined. Both thermal stability and stability, toward the denaturing agents guanidinium chloride and urea, of free and immobilized enzymes were compared next. The thermal stability of the immobilized enzyme is significantly improved in comparison with free HRP. The catalytic kinetics is slowed down notably, but V max is much more robust to heat than the free enzyme. The stability resistance of the enzyme toward the denaturing agents depends on the chemical nature of the denaturing agents and interactions between enzyme and silica nanopore walls. Guanidinium chloride showed similar attenuation of the catalytic activity of immobilized and free enzyme. In contrast, the immobilized HRP was much more resistant to urea than the free enzyme. The different behavior of free and immobilized enzyme is most likely due to different hydrogen bonding of water and increased hydration strength of the protein inside the nanopores.

Many key chemical and biochemical reactions, particularly in living cells, take place in confined space at the mesoscopic scale. Toward understanding of physicochemical nature of biomacromolecules confined in nanoscale space, in this work we have elucidated fluorescence effects of a light harvesting complex LH2 in nanoscale chemical environments. Mesoporous silicas (SBA-15 family) with different shapes and pore sizes were synthesized and used to create nanoscale biomimetic environments for molecular confinement of LH2. A combination of UV−vis absorption, wide-field fluorescence microscopy, and in situ ellipsometry supports that the LH2 complexes are located inside the silica nanopores. Systematic fluorescence effects were observed and depend on degree of space confinement. In particular, the temperature dependence of the steady-state fluorescence spectra was analyzed in detail using condensed matter band shape theories. Systematic electronicvibrational coupling differences in the LH2 transitions between the free and confined states are found, most likely responsible for the fluorescence effects experimentally observed.

Peritoneal carcinomatosis results from dissemination of solid tumors in the peritoneal cavity, and is a common site of metastasis in patients with carcinomas of gastrointestinal or gynecological origin. Peritoneal carcinomatosis treatment is challenging as poorly vascularized, disseminated peritoneal micro-tumors are shielded from systemic anticancer drugs and drive tumor regrowth. Here, we describe the identification and validation of a tumor homing peptide CKRDLSRRC (IP3), which upon intraperitoneal administration delivers payloads to peritoneal metastases. IP3 peptide was identified by in vivo phage display on a mouse model of peritoneal carcinomatosis of gastric origin (MKN-45P), using high-throughput sequencing of the peptide-encoding region of phage genome as a readout. The IP3 peptide contains a hyaluronan-binding motif, and fluorescein-labeled IP3 peptide bound to immobilized hyaluronan in vitro. After intraperitoneal administration in mice bearing peritoneal metastases of gastric and colon origin, IP3 peptide homed robustly to macrophage-rich regions in peritoneal tumors, including poorly vascularized micro-tumors. Finally, we show that IP3 functionalization conferred silver nanoparticles the ability to home to peritoneal tumors of gastric and colonic origin, suggesting that it could facilitate targeted delivery of nanoscale payloads to peritoneal tumors. Collectively, our study suggests that the IP3 peptide has potential applications for targeting drugs, nanoparticles, and imaging agents to peritoneal tumors.

Synopsis: structural and catalytic features of a complex enzyme galactose oxidase confined in nanoscale chemical environments were investigated to show the catalytic efficiency of the enzyme depending on both the degree of space confinement and immobilization method.