Correlation of [RuCl3(dppb)(VPy)] cytotoxicity with its effects on the cell membranes: an investigation using Langmuir monolayers as membrane models.

One of the major challenges in drug design is to identify compounds with potential toxicity toward target cells, preferably with molecular-level understanding of their mode of action. In this study, the antitumor property of a ruthenium complex, mer-[RuCl3(dppb)(VPy)] (dppb = 1,4-bis(diphenylphosphine)butane and VPy = 4-vinylpyridine) (RuVPy), was analyzed. Results showed that this compound led to a mortality rate of 50% of HEp-2 cell with 120 ± 10 µmol L(-1), indicating its high toxicity. Then, to prove if its mode of action is associated with its interaction with cell membranes, Langmuir monolayers were used as a membrane model. RuVPy had a strong effect on the surface pressure isotherms, especially on the elastic properties of both the zwitterionic dipalmitoylphosphatidylcholine (DPPC) and the negatively charged dipalmitoylphosphatidylglycerol (DPPG) phospholipids. These data were confirmed by polarization-modulated infrared reflection-absorption spectroscopy (PM-IRRAS). In addition, interactions between the positive group from RuVPy and the phosphate group from the phospholipids were corroborated by density functional theory (DFT) calculations, allowing the determination of the Ru complex orientation at the air-water interface. Although possible contributions from receptors or other cell components cannot be discarded, the results reported here represent evidence for significant effects on the cell membranes which are probably associated with the high toxicity of RuVPy.

On the use of surface neutron-gamma gauges to estimate soil water content.

Surface neutron-gamma gauges are handy instruments to measure soil water contents and bulk densities of surface layers. Although available for some decades, their optimal use is still not well established. This study is a contribution to improve their use, mainly in relation to calibration, and of the effect of soil dry bulk density on soil water content measurements.

Aggregation phenomena in the complexes of iron tetraphenylporphine sulfonate with bovine serum albumin.

Binding of Fe(III) meso-tetrakis(p-sulfonatophenyl)-porphyrin (FeTPPS4) to bovine serum albumin (BSA) was studied by UV-VIS absorption, fluorescence quenching, circular dichroism, 1H NMR, and ESR. At excess of BSA, the bound form of FeTPPS4 is a high-spin monomer exhibiting a Soret band at 417 nm, a broad NMR peak at 10.3 ppm, an ESR signal at g = 5.7-5.9, and a strong enhancement of magnetic relaxation of water protons. In the intermediate concentration range, a formation of nonparamagnetic bound aggregates of FeTPPS4 occurs (up to 10-15 molecules at pH 6.0) with a Soret band at 414 nm and NMR peaks at 7.0, 8.1, and 12.7 ppm. In the physiologic pH range, BSA binds the monomeric form of FeTPPS4 with an association constant of about 10(8) M-1, the affinity to oxo-dimers in solution being much lower. BSA itself is also subject to aggregation with an average aggregation number of 4-8 in the physiological pH range. It is assumed that aggregation phenomena may play an important role, both in the relaxation efficiency of metalloporphyrins as MRI contrast agents and in the blood transport of porphyrin drugs by albumins.

Spectroscopic studies of the met form of the extracellular hemoglobin from Glossoscolex paulistus.

Sephadex G-200 chromatography of the extracellular hemoglobin from the giant earthworm G. paulistus in the met form presents a single peak at pH 7.0 and two peaks at pH 9.0 as a result of alkaline dissociation. SDS-PAGE shows that the polypeptide chains are very similar to those observed for the oxy form and the two peaks at pH 9.0 correspond to the trimer contaminated by linkers and monomers which seems to be quite pure. The aquomet acid form is stable as an oligomer of molecular mass 3.1 x 10(6) Da only in a narrow pH range around neutrality. Increasing the pH above 7.5 leads to an irreversible transition from aquomet to hemichrome I which is the low-spin bis-imidazole complex. At pHs above 9.5-10.0 a second reversible transition takes place from hemichrome I to hemichrome II, a high-spin complex which is associated with the weakening and possible disruption of the proximal Fe--N histidine bond. Thus, increase in pH above 8.0 induces changes in the heme pocket that involve both the distal and proximal sides of the heme. EPR measurements show a very sharp decrease of the aquomet high-spin signal in the range of pH 7.0-8.0 and a very small low-spin signal even at liquid helium temperatures. The transition to hemichrome I is also accompanied by the loss of heme optical activity monitored by CD, which is consistent with the weakening of heme--globin interaction. Hemichrome I in the presence of cyanide gives the typical cyanometHb derivative which has a transition to a hemichrome at much higher pHs. This observation suggests that the dissociation of the oligomer in alkaline medium as well as the stability of the heme on the proximal side, depend both upon the ligand present at the sixth coordination position on the distal side. Hence, we believe that hemi(hemo)chrome formation in G. paulistus Hb and other invertebrate hemoglobins is a common phenomenon, not associated with protein denaturation, which may provide a fine tuning mechanism to control subunit interactions through changes in the distal side of the heme pocket.

H NMR and electronic absorption spectroscopy of paramagnetic water-soluble meso-tetraarylsubstituted cationic and anionic metalloporphyrins.

The ionization, mu-oxo-dimerization and axial ligation equilibria of free bases, iron(III) and manganese(III) derivatives of meso-tetrakis(p-sulfonatophenyl)porphyrin (TPPS4) and meso-tetrakis(4-N-methyl-pyridiniumyl)porphyrin (TMPyP) in aqueous solution are studied by 1H NMR and electronic absorption spectroscopy. At physiological pH, Fe(III) complexes of TMPyP and TPPS4 exist predominantly as dimers and may undergo transition to low spin species upon binding to biomolecules, whereas Mn(III) complexes are essentially monomeric. Dicyano and bis-imidazole complexes of FeTMPyP and FeTPPS4 are low spin monomer adducts in the pH range 2.0 to 11.2. No low spin dimeric complexes were found. The low spin monocyano and high spin mono-imidazole complexes of FeTMPyP are formed in acidic and alkaline media, respectively. T1-relaxation enhancement of water protons at 200 MHz induced by FeTPPS4 falls dramatically in the sequence high spin >> dimeric > low spin form.

Binding of manganese and iron tetraphenylporphine sulfonates to albumin is relevant to their contrast properties.

The interaction of Fe(III) and Mn(III) complexes of TPPS4 with bovine serum albumin (BSA) was studied by T1 relaxation measurements of water protons and high resolution 1H NMR of the porphyrin moieties. At excess of BSA, both metalloporphyrins bind to BSA as the high spin monomers. The relaxivity of bound MnTPPS4 is significantly higher as compared to the free form in solution. When metalloporphyrins are in excess, they aggregate at the BSA surface, up to two MnTPPS4, and up to 10-15 FeTPPS4 units per BSA globule. Bound aggregates are unable to enhance magnetic relaxation of water protons due to the antiferromagnetic coupling between metal ions in the aggregates. Therefore, the dose-effect dependences for metalloporphyrins in the range of metalloporphyrin/BSA ratio of 0 to 25 at the constant BSA concentration at pH 7.4 are characterized by a local maximum at about 2 for MnTPPS4, and a global maximum at about 3 for FeTPPS4, MnTPPS4 complex is more effective than FeTPPS4 in the whole concentration range. It is suggested that the difference in binding and aggregation properties of metalloporphyrins may be relevant to their relaxation efficiency in vivo, blood transport, and biodistribution.

Inhibition of ischemic brain edema formation by post-ischemic administration of a prostaglandin oligomer.

We studied the efficacy of an oligomeric derivative of prostaglandin E1 in protecting the rat brain against focal ischemia. The degree of ischemic damage was evaluated from three parameters, namely, the degree of edema formation, reduction of motor performance, and memory disturbance as measured by a passive avoidance test. The pre-ischemic administration of the drug (6 mg/kg i.p.) had some effects, but the differences were not significant. The post-ischemic administration (6 mg/kg i.p.) produced significant improvement in all three parameters. The increase of water content of the ischemic hemisphere was reduced (p less than 0.05); the total motor score was improved (p less than 0.01); and the memory disturbance as estimated by the passive avoidance test was reduced (p less than 0.01). A possible mechanism of protection is discussed.