Vancomycin covalently bonded to titanium alloy prevents bacterial colonization.

Periprosthetic infection is a devastating consequence of implant insertion and can arise from hematogenous sources or surgical contamination. Microbes can preferentially colonize the implant surface and, by forming a biofilm, escape immune surveillance. We hypothesized that if an antibiotic can be tethered to a titanium alloy (Ti) surface, it will inhibit bacterial colonization, prevent biofilm formation, and avert late-stage infection. To test this hypothesis, a Ti rod was covalently derivatized with vancomycin. Reaction efficiencies were evaluated by colorimetric and spectrophotometric measurements. The vancomycin-modified surface was stable in aqueous solutions over extended time periods and maintained antibiotic coverage, even after press-fit insertion into a cadaverous rat femora. When evaluated using fluorescently labeled bacteria, or by direct colony counts, the surface-bound antibiotic prevented bacterial colonization in vitro after: (1) exposure to high levels of S. aureus; (2) extended incubation in physiological buffers; and (3) repeated bacterial challenges. Importantly, whereas the vancomycin-derivitized pins prevented bacterial colonization, S. aureus adhered to control pins, even in the presence of concentrations of vancomycin that exceeded the strain MIC. These results demonstrate that we have effectively engineered a stable, bactericidal Ti surface. This new surface holds great promise in terms of mitigating or preventing periprosthetic infection.

Covalent bonding of vancomycin to Ti6Al4V alloy pins provides long-term inhibition of Staphylococcus aureus colonization.

Self-protecting Ti6Al4V alloy pins were prepared by covalent bonding of bis(ethylene glycol) linkers, then vancomycin to the oxidized, aminopropylated Ti6Al4V alloy surface. Fluorescence modification-enabled estimation of yields of free amines on the metallic surface monolayer at each reaction step. The vancomycin-protected Ti6Al4V pins were not colonized by Staphylococcus aureus, even after 44days storage in physiological buffer. These results provide a basis for testing self-protection against S. aureus colonization in animal models.

A protocol for high frequency regeneration through nodal explant cultures and ex vitro rooting of Plumbago rosea L.

A rapid clonal multiplication scheme comprising direct multiple shoot initiation and downsizing of the node with buds proliferated upon during subculture was developed for Plumbago rosea. Sixty five per cent of the nodes (approximately 2.0 cm) dissected out of young shoots from field grown plants and cultured in MS agar medium containing 3% sucrose and 15.4 microM BAP remained contamination free and responded at 95% rate with callusing at basal cut end and axillary bud break in 5 days followed by the formation of 2.41 +/- 0.14 shoots of 0.87 +/- 0.14 cm length in 3 weeks. Though differences in frequency and number of buds formed between nodes of 1-5 positions from the young shoots was negligible, the shoots emanated from the youngest node were shorter (0.92 +/- 0.19 cm) than those (2.3 +/- 0.50 cm) of the mature 5th node. Synergistic influence of BAP and auxins on caulogenesis was absent. Bud emergence in shorter (approximately 0.5 cm) nodes was delayed up to 3 weeks and extensive callus proliferation from the cut basal end overlapped the 8.2 +/- 0.37 axillary shoots/buds formed after 7 weeks. Reduction in the size (downsized) of the 2.0 cm node with buds to 1.0 cm by dissecting out the basal internodal segment having the callus and subculture of them (approximately 1.0 cm) with buds in contact with the medium for 3 weeks contributed to maximum multiplication of 42.1 +/- 5.40 shoot buds. Division of the shoot cluster and transfer of 2-3 shoots (0.5-1.5 cm) in a clump to MS basal liquid medium induced elongation of the shoots to 4.1 +/- 0.18 cm in 2 weeks. Shoots of 3.0-4.2 cm length were rooted within 3 weeks at 100% efficiency in vitro or ex vitro without hardening. In vitro rhizogenesis in presence of 0.49 microM IBA is recommended for enhanced rooting and high yield of commercially important tuberous roots during cultivation in the field.

Vancomycin covalently bonded to titanium beads kills Staphylococcus aureus.

Periprosthetic infections are life-threatening complications that occur in about 6% of medical device insertions. Stringent sterile techniques have reduced the incidence of infections, but many implant patients are at high risk for infection, especially the elderly, diabetic, and immune compromised. Moreover, because of low vascularity at the site of the new implant, antibiotic prophylaxis is often not effective. To address this problem, we designed a covalent modification to titanium implant surfaces to render them bactericidal. Specifically, we aminopropylated titanium, a widely used implant material and extended a tether by solid phase coupling of ethylene glycol linkers, followed by solid phase coupling of vancomycin. Vancomycin covalently attached to titanium still bound soluble bacterial peptidoglycan, reduced Staphylococcus aureus colony-forming units by 88% +/- 16% over 2 hr, and retained antibacterial activity upon a repeated challenge.

Chlamydocin-hydroxamic acid analogues as histone deacetylase inhibitors.

Chlamydocin-hydroxamic acid analogues were designed and synthesized as histone deacetylase (HDAC) inhibitors based on the structure and HDAC inhibitory activity of chlamydocin and trichostatin A. Chlamydocin is a cyclic tetrapeptide containing an epoxyketone moiety in the side chain that makes it an irreversible inhibitor of HDAC. We replaced the epoxyketone moiety of chlamydocin with hydroxamic acid to design potent and reversible inhibitors of HDAC. In addition, a number of amino-cycloalkanecarboxylic acids (Acc) are introduced instead of the simple amino-isobutric acid (Aib) for a variety of the series of chlamydocin analogues. The compounds synthesized were tested for HDAC inhibitory activity and the results showed that many of them are potent inhibitors of HDAC. The replacement of Aib residue of chlamydocin with an aromatic amino acid enhances the in vivo and in vitro inhibitory activity. We have carried out circular dichroism and molecular modeling studies on chlamydocin-hydroxamic acid analogue and compared it with the solution structure of chlamydocin.

Novel histone deacetylase inhibitors: cyclic tetrapeptide with trifluoromethyl and pentafluoroethyl ketones.

Cyclic tetrapeptides containing trifluoromethyl and pentafluoroethyl ketone as zinc binding functional group were synthesized as potent HDAC inhibitors. Evaluation by human HDAC inhibition assay and p21 promoter assay showed that these inhibitors are promising anticancer agents.

In vitro mass multiplication of Ophiorrhiza mungo Linn.

A protocol for in vitro mass multiplication of plants through seedling (shoot) cultures was established for Ophiorrhiza mungo. Maximum number of adventitious shoots per shoot culture (10.4 +/- 1.72) was initiated on MS solid medium supplemented with BAP (2.22 microM) after 3 weeks. Shoots were further multiplied (12.8 +/- 2.8) through subculture of intact shoots and reculture of nodal segments of aseptic shoots (6.5 +/- 0.94) in MS solid medium containing BAP (0.89 microM). Shoot elongation (1.27 +/- 0.12 cm) was achieved in the medium containing GA3 (1.44 microM) in two weeks. Rooting was favoured in basal agar medium supplemented with IBA (12.3 microM) plus NAA (1.07 microM). The plants were successfully established (100%) in the pots containing sand and top soil (1:1) mixture in a period of two weeks.

Synthesis and histone deacetylase inhibitory activity of cyclic tetrapeptides containing a retrohydroxamate as zinc ligand.

Cyclic tetrapeptide retrohydroxamic acids were prepared as histone deacetylase (HDAC) inhibitors and evaluated the inhibitory activity and found that they have potential as anticancer drugs.

Toward an HDAC6 inhibitor: synthesis and conformational analysis of cyclic hexapeptide hydroxamic acid designed from alpha-tubulin sequence.

A cyclic hexapeptide hydroxamic acid inhibitor for HDAC6 has been designed and synthesized on the basis of the facts that alpha-tubulin is the substrate of HDAC6 and of the excellent inhibitory activity of cyclic tetrapeptide hydroxamic acids (CHAPs) for HDACs. Unexpectedly, cyclic hexapeptide hydroxamic acid showed very low HDAC inhibitory activity. To explain the low activity, we have carried out conformation analysis and compared it to the crystal structure of alpha-tubulin. The conformation around the acetylated lysine of the cyclic hexapeptide substrate or the aminosuberate hydroxamic acid [Asu(NHOH)] of cyclic hexapeptide inhibitor is different from that around alpha-tubulin's lysine-40. The difference in the conformation seems to cause some steric hindrance at the capping site resulting in poor binding capacity.

Cyclic tetrapeptides bearing a sulfhydryl group potently inhibit histone deacetylases.

New inhibitors of histone deacetylase (HDAC) containing a sulfhydryl group were designed on the basis of the corresponding hydroxamic acid (CHAP31) and FK228. Their disulfide dimers and hybrids exhibited potent HDAC inhibitory activity in vivo with potential as anticancer prodrugs. [structure: see text]