Anticaries potential of commercial fluoride rinses as determined by fluoridation and remineralization efficiency.

OBJECTIVE: The objective of this work was to compare the anticaries potential of several currently marketed fluoride-containing mouthrinse products using two in vitro approaches: 1) fluoride uptake studies of demineralized human enamel samples after exposure to rinse products; and 2) microhardness studies of sound enamel samples after exposure to the rinse products and demineralizing agents. METHODS: Four currently marketed rinse products, formulated at 100 ppm F, were evaluated in fluoride uptake studies relative to a negative (water) rinse control (Study 1). The same rinse products were evaluated in microhardness studies (Study 2) against a positive control, ACT Anticavity rinse, which is formulated with 225 ppm F and carries the ADA Seal of Acceptance as an effective anticavity mouthrinse. Test products included ACT Total Care rinse (pH = 6.34), Listerine Total Care rinse (pH = 3.57), Crest Pro-Health for Me rinse (pH = 3.33), and Crest Pro-Health Complete rinse (pH = 3.43). RESULTS: Study 1-Samples treated with any of the fluoride-containing rinses showed significantly higher (p < 0.05) levels of fluoride uptake than the negative (water) control. Two of the products (Crest Pro-Health for Me and Crest Pro-Health Complete) showed significantly higher (p < 0.05) levels of fluoride uptake into demineralized enamel than the other marketed rinses (Listerine Total Care and ACT Total Care). Study 2-Samples treated with the same two rinse products (Crest Pro-Health For Me and Crest Pro-Health Complete) showed significantly lower mineral loss than the other rinse products, as well as the positive control. CONCLUSION: Results of these in vitro studies indicate that the Crest mouthrinse products evaluated here are capable of providing significantly better fluoridation of demineralized enamel, as well as significantly better protection against the initiation and progression of demineralization, compared to the other marketed fluoride-containing mouthrinse products tested.

Synthesis and in vitro evaluation of human FP-receptor selective prostaglandin analogues.

The in vitro evaluation of a series of saturated prostaglandins revealed that compounds with omega chain aromatic rings retain nanomolar potency for the human prostaglandin F receptor (hFP receptor), exemplified by compound 8. In contrast, the double bonds are required for activity in the series with an acyclic omega chain as in PGF2alpha.

Use of short high-performance liquid chromatography columns and tandem-mass spectrometry for the rapid analysis of a prostaglandin analog, fluprostenol, in rat plasma.

A short reversed-phase HPLC column and a tandem mass spectrometer were used to develop a stable-isotope-dilution assay for the rapid and sensitive analysis of fluprostenol, a prostaglandin analog, in rat plasma. A Waters Symmetry ODS column (2.1x10 mm) afforded rapid isocratic elution of fluprostenol (t(R)=40 s) but still provided a relatively large k' value of 4. The use of tandem mass spectrometry allowed the interference-free detection of fluprostenol under the rapid elution conditions, with a limit of quantitation of 25 pg ml(-1) fluprostenol, using 0.2 ml plasma sample volumes. The method was linear over three orders of magnitude, yielded accurate and precise results and allowed the pharmacokinetic profile of fluprostenol to be defined following intravenous administration in rats.

Design and synthesis of 13,14-dihydro prostaglandin F(1alpha) analogues as potent and selective ligands for the human FP receptor.

The in vitro evaluation of a new class of potential bone anabolic agents for the treatment of osteoporosis is described. These compounds are potent and selective ligands for the human prostaglandin F receptor (hFP receptor). The compounds lack the olefin unsaturation required for potency in the natural ligand PGF(2)(alpha) yet retain binding affinity for the hFP receptor in the nanomolar to micromolar range. Removal of the alkenes also results in a better selectivity ratio for the hFP receptor over the other prostaglandin receptors tested. A rationale for the selectivity differences of various analogues, based on ligand docking experiments to a putative hFP receptor model, is also described.

Characterization of lanthanide ion binding to the EF-hand protein S100 beta by luminescence spectroscopy.

S100 beta is a member of a group of low-molecular weight acidic calcium binding proteins widely distributed in the vertebrate nervous system containing two helix-loop-helix calcium binding motifs (sites I and II). In addition, S100 beta also has auxiliary Zn2+ binding sites that are distinct from the Ca2+ binding sites. Luminescence spectroscopy using Eu3+ and Tb3+ as spectroscopic probes for Ca2+ is used to characterize the Ca2+ binding sites of this protein. Eu3+-bound S100 beta shows two distinct Eu3+ binding environments from both the excitation spectrum and Eu3+ excited state lifetimes. Eu3+ bound to the classical EF hand site II has a Kd of 660 +/- 20 nM, whereas the dissociation constant for the pseudo-EF hand site I is significantly weaker. Lifetimes in H2O and D2O lead to the finding that there are four water molecules coordinated to the Eu3+ in the weakly binding site I and two water molecules to the tightly binding site II. Site II in S100 beta expectedly is very similar to high-affinity Ln3+ binding domains I and II in calmodulin. Eu3+ luminescence experiments with Zn2+-loaded S100 beta show that the lifetime for Eu3+ in site I in Zn2+-loaded S100 beta is significantly different than that in the absence of Zn2+. Tyrosine-17-sensitized Tb3+ luminescence experiments indicate that the Tb3+ occupying the proximal weaker binding site I is sensitized, whereas Tb3+ in site II is not. The distance between sites I and II (15.0 +/- 0.4 A) in S100 beta was determined from Forster-type energy transfer in D2O solutions containing bound Eu3+ donor and Nd3+ acceptor ions. For Zn2+-S100 beta, the intersite distance is reduced to 13 +/- 0.3 A. Location of histidine-15 close to pseudo-EF site I suggests that Zn2+ binding likely changes the conformation of this site, causing a reduction of the intersite distance by approximately 2 A.

Solution structure of rat apo-S100B(beta beta) as determined by NMR spectroscopy.

S100B(beta beta), a member of the S100 protein family, is a Ca(2+)-binding protein with noncovalent interactions at its dimer interface. Each apo-S100 beta subunit (91 residues) has four alpha-helices and a small antiparallel beta-sheet, consistent with two predicted helix-loop-helix Ca(2+)-binding domains known as EF-hands [Amburgey et al. (1995) J. Biomol. NMR 6, 171-179]. The three-dimensional solution structure of apo-S100B(beta beta) from rat has been determined using 2672 distance (14.7 per residue) and 88 dihedral angle restraints derived from multidimensional nuclear magnetic resonance spectroscopy. Apo-S100B (beta beta) is found to be globular and compact with an extensive hydrophobic core and a highly charged surface, consistent with its high solubility. At the symmetric dimer interface, 172 intermolecular nuclear Overhauser effect correlations (NOEs) define the antiparallel alignment of helix I with I' and of helix IV with IV'. The perpendicular association of these pairs of antiparallel helices forms an X-type four-helical bundle at the dimer interface. Whereas, the four helices within each apo-S100 beta subunit adopt a unicornate-type four-helix bundle, with helix I protruding from the parallel bundle of helices II, III, and IV. Accordingly, the orientation of helix III relative to helices I, II, and IV in each subunit differs significantly from that known for other Ca(2+)-binding proteins. Indeed, the interhelical angle (omega) observed in the C-terminal EF-hand of apo-S100 beta is -142 degrees, whereas omega ranges from 118 degrees to 145 degrees in the apo state and from 84 degrees to 128 degrees in the Ca(2+)-bound state for the EF-hands of calbindin D9k, calcyclin, and calmodulin. Thus, a significant conformational change in the C-terminal EF-hand would be required for it to adopt a structure typical of the Ca(2+)-bound state, which could readily explain the dramatic spectral effects observed upon the addition of Ca2+ to apo-S100B(beta beta).

1H, 13C and 15N NMR assignments and solution secondary structure of rat Apo-S100 beta.

The 1H, 13C and 15N NMR assignments of the backbone and side-chain resonances of rat S100 beta were made at pH 6.5 and 37 degrees C using heteronuclear multidimensional NMR spectroscopy. Analysis of the NOE correlations, together with amide exchange rate and 1H alpha, 13C alpha and 13C beta chemical shift data, provided extensive secondary structural information. Thus, the secondary structure of S100 beta was determined to comprise four helices (Leu3-Ser18, helix I; Lys29-Leu40, helix II; Gln50-Glu62, helix III; and Phe70-Ala83, helix IV), four loops (Gly19-His25, loop I; Ser41-Glu49, loop II; Asp63-Gly66, loop III; and Cys84-Glu91, loop IV) and two beta-strands (Lys26-Lys28, beta-strand I and Glu67-Asp69, beta-strand II). The beta-strands were found to align in an antiparallel manner to form a very small beta-sheet. This secondary structure is consistent with predictions that S100 beta contains two 'helix-loop-helix' Ca(2+)-binding motifs known as EF-hands. The alignment of the beta-sheet, which brings the two EF-hand domains of S100 beta into close proximity, is similar to that of several other Ca(2+)-ion-binding proteins.