Masahiko Chikuma

Determination of hydrogen peroxide with N,N-diethylaniline and 4-aminoantipyrine by use of an anion-exchange resin modified with manganese-tetrakis(sulphophenyl)porphine, as a substitute for peroxidase

Amberlite IRA 900 anion-exchange resin modified with manganese-tetrakis(sulphophenyl)-porphine has been used as a catalyst instead of peroxidase for the determination of hydrogen peroxide by the reaction 2H(2)O(2) + N,N-diethylaniline + 4-aminoantipyrine (catalyst)--> quinonoid dye (lambda(max) 550 nm) + 4H(2)O. The apparent molar absorptivity for hydrogen peroxide was 1.1 x 10(4) 1.mole(-1).cm(-1), coefficient of variation 0.7%. This value is approximately 84% of that obtained by the use of peroxidase as catalyst. Similar conditions to those in the enzymatic reaction were suitable for use of the modified resin as catalyst, and the results show it to be a good substitute for peroxidase in this reaction system.

The phosphate clamp: a small and independent motif for nucleic acid backbone recognition

The 1.7 Å X-ray crystal structure of the B-DNA dodecamer, [d(CGCGAATTCGCG)]2 (DDD)-bound non-covalently to a platinum(II) complex, [{Pt(NH3)3}2-µ-{trans-Pt(NH3)2(NH2(CH2)6NH2)2}](NO3)6 (1, TriplatinNC-A,) shows the trinuclear cation extended along the phosphate backbone and bridging the minor groove. The square planar tetra-am(m)ine Pt(II) units form bidentate N-O-N complexes with OP atoms, in a Phosphate Clamp motif. The geometry is conserved and the interaction prefers O2P over O1P atoms (frequency of interaction is O2P > O1P, base and sugar oxygens > N). The binding mode is very similar to that reported for the DDD and [{trans-Pt(NH3)2(NH2(CH2)6(NH3+)}2-µ-{trans-Pt(NH3)2(NH2(CH2)6NH2)2}](NO3)8 (3, TriplatinNC), which exhibits in vivo anti-tumour activity. In the present case, only three sets of Phosphate Clamps were found because one of the three Pt(II) coordination spheres was not clearly observed and was characterized as a bare Pt2+ ion. Based on the electron density, the relative occupancy of DDD and the sum of three Pt(II) atoms in the DDD-1 complex was 1:1.69, whereas the ratio for DDD-2 was 1:2.85, almost the mixing ratio in the crystallization drop. The high repetition and geometric regularity of the motif suggests that it can be developed as a modular nucleic acid binding device with general utility.

A Tetrazolato-Bridged Dinuclear Platinum(II) Complex Exhibits Markedly High in vivo Antitumor Activity against Pancreatic Cancer

Some platinum(II) coordination complexes are effective anticancer agents. cis-Diamminedichloridoplatinum(II) (cisplatin), a mononuclear platinum(II) complex, is one of the most commonly used anticancer drugs. Although platinum-based chemotherapy can cause serious side effects, its efficacy has prompted the design and synthesis of next-generation anticancer platinum(II) drugs which are effective against cancers that are typically resistant to chemotherapy, such as lung cancer, pancreatic cancer, and platinum-refractory cancer. 3] Lung cancer is the leading cause of cancer deaths worldwide, and non-small-cell lung cancer (NSCLC) accounts for 80 % of lung cancers. Pancreatic cancer remains the fourth leading cause of cancer-related deaths in the United States. Clinical platinum-based drugs show antitumor efficacy by forming Pt–DNA adducts. We and others have reported that azolato-bridged dinuclear Pt complexes such as [{cis-Pt(NH3)2}2(m-OH)(m-pyrazolato)](NO3)2 (1) and [{cis-Pt(NH3)2}2(mOH)(m-1,2,3-triazolato-N1,N2)](NO3)2 (2), interact with DNA through a mechanism different from that of cisplatin and exhibit much higher in vitro cytotoxicity than cisplatin. The chemical structures of 1 and 2 are shown in Figure 1. To further expand drug discovery, we designed two new tetrazolatobridged complexes, [{cis-Pt(NH3)2}2(m-OH)(m-tetrazolatoN1,N2)](ClO4)2 (3) and [{cis-Pt(NH3)2}2(m-OH)(m-tetrazolatoN2,N3)](ClO4)2 (4), which are structural isomers (Figure 1). Herein we report their synthesis, characterization, in vitro cytotoxicity, and preliminary in vivo antitumor efficacy. Complexes 3 and 4 were synthesized by using a modified protocol for azolato-bridged complexes, as previously described (Scheme 1). A slight excess of tetrazole was added to an aqueous solution of the starting material, [cis-Pt(NH3)2(mOH)]2(NO3)2, which was then incubated at 40 8C in the dark to yield 3 and 4 at a molar ratio of 6.5:3.5 (see Supporting Information). This preparation ratio likely resulted from the higher nucleophilicity of N1 or N4 bound to one carbon and one nitrogen atom, compared with N2 or N3 bound to two nitrogen atoms. The two structural isomers were separated and purified by reversed-phase liquid chromatography and characterized by H, C, and Pt NMR spectroscopy and ESI mass spectrometry. For 3, two Pt NMR chemical shifts appeared at 2127 and 2177 ppm, reflecting two slightly different Pt[N3O] environments, because the tetrazolato bridge is arranged in an asymmetric fashion; there is N1,N2 coordination of Pt atoms. In contrast, for 4, a single Pt NMR chemical shift appears at about 2180 ppm, confirming the Pt[N3O] environment and the symmetric structure of the complexes on the tetrazolato bridge; there is N2,N3 coordination of Pt atoms. We tested the cytotoxicity of our azolato-bridged dinuclear Pt complexes, along with cisplatin for comparison, on the H460 human NSCLC cell line; the IC50 values are listed in Table 1. Complex 1 showed no activity at concentrations Figure 1. Structures of (1) [{cis-Pt(NH3)2}2(m-OH)(m-pyrazolato-N1,N2)] 2 + , (2) [{cis-Pt(NH3)2}2(m-OH)(m-1,2,3-triazolato-N1,N2)] 2+ , (3) [{cis-Pt(NH3)2}2(m-OH)(mtetrazolato-N1,N2)] + , and (4) [{cis-Pt(NH3)2}2(m-OH)(m-tetrazolato-N2,N3)] 2 +

Selective sorption of fluoride ions by anion-exchange resin modified with alizarin fluorine blue-praseodymium(III) complex

Abstract A new functional resin for selective sorption of fluoride ions was prepared from an anion-exchange resin (Amberlite IRA 400) and a praseodymium(III) complex of alizarin fluorine blue (AFB). The AFB-Pr(III) complex was immobilized by both ion-exchange and nonelectrostatic interaction between AFB molecules and the matrix of the anion-exchange resin. The AFB-Pr(III) complex formed a ternary complex with fluoride ions in the resin. The resin modified with the AFB-Pr(III) complex (AFB-Pr resin) had specific and nonspecific binding sites for sorption of fluoride ions. The former were ligand-exchange sites where fluoride ions replace water molecules coordinated to praseodymium(III) in the resin, and the latter were residual ion-exchange sites where fluoride ions replace chloride ions of quarternary ammonium chloride groups. Fluoride ions were sorbed strongly and specifically in the presence of electrolytes by the former sites. The sorption capacity for fluoride ions was about 0.5 mg/g resin in the presence of 0.1 M sodium chloride when 0.028 mmol of AFB-Pr(III) complex was immobilized per gram of the anion-exchange resin. The effects of pH, buffer, acetone, and foreign ions on fluoride uptake were examined by a batch method. Fluoride uptake decreased with increasing pH. The addition of acetone enhanced fluoride uptake at both binding sites. Interference from the cations and anions studied was negligible except for aluminum(III). Fluoride sorbed on AFB-Pr resin was removed with a dilute sodium hydroxide solution.

Highly efficient DNA compaction mediated by an in vivo antitumor-active tetrazolato-bridged dinuclear platinum(II) complex.

We investigated the effects of antitumor-active tetrazolato-bridged dinuclear platinum(II) complexes [{cis-Pt(NH(3))(2)}(2)(μ-OH)(μ-tetrazolato-N(1),N(2))](2+) (1) and [{cis-Pt(NH(3))(2)}(2)(μ-OH)(μ-tetrazolato-N(2),N(3))](2+) (2) on the higher-order structure of a large DNA molecule (T4 phage DNA, 166 kbp) in aqueous solution through single-molecule observation by fluorescence microscopy. Complexes 1 and 2 cause irreversible compaction of DNA through an intermediate state in which coil and compact parts coexist in a single DNA molecule. The potency of compaction is in the order 2 > 1 ≫ cisplatin. Transmission electron microscopic observation showed that both complexes collapsed DNA into an irregularly packed structure. Circular dichroism measurements revealed that the dinuclear platinum(II) complexes change the secondary structure of DNA from the B to C form. These characteristics of platinum(II) complexes are markedly different from those of the usual condensing agents such as spermidine(3+) and [Co(III)(NH(3))(6)](3+). The ability to cause DNA compaction by the platinum(II) complexes is discussed in relation to their potent antitumor activity.

Determination of selenium(IV) and other forms of selenium dissolved in sea water by anion-exchange resin loaded with sulfonic acid derivative of bismuthiol-II and hydride generation atomic-absorption spectrometry

SummaryA sulfonic acid derivative of bismuthiol-II (bisIIS) was synthesized from 4-hydrazinobenzene sulfonic acid and carbon disulfide. Selenium(IV) was adsorbed selectively and quantitatively on the anion-exchange resin loaded with bis-IIS. Selenium adsorbed on the resin was eluted by the use of penicillamine and determined by hydride generation atomic absorption spectrometry (hydride generation/AAS). Selenium(VI) and other forms of selenium, which were not adsorbed onto the resin, were collected on the resin after digestion with nitric acid followed by reduction with hydrochloric acid. Separative preconcentration of selenium(IV), selenium(VI) and other forms of selenium in 0.5 mol/l sodium chloride could be carried out successfully by the proposed procedures. However, in the case of estuarial sea water containing a large quantity of organic substances, selenium(IV) could not be separated, because organic substances interfered with the reduction of selenium(VI) to selenium(IV) by the use of hydrochloric acid. Selenium(IV) and total amount of selenium(VI) and other forms of selenium dissolved in polluted sea water samples were determined by the proposed procedures.

Synthesis of antitumor azolato-bridged dinuclear platinum(II) complexes with in vivo antitumor efficacy and unique in vitro cytotoxicity profiles

We synthesised four tetrazolato-bridged dinuclear Pt(ii) complexes, [{cis-Pt(NH3)2}2(μ-OH)(μ-5-R-tetrazolato-N2,N3)](n+), where R is CH3 (1), C6H5 (2), CH2COOC2H5 (3), or CH2COO(-) (4) and n = 2 (1-3) or 1 (4). Their structures were characterised by (1)H, (13)C, and (195)Pt NMR spectroscopy, mass spectrometry, and elemental analysis, and the crystal structure of 1 was determined by X-ray crystallography. The cytotoxicities of the complexes to human non-small-cell lung cancer (NSCLC) cell lines sensitive and resistant to cisplatin were assayed. Complex 1 was more cytotoxic than cisplatin in both PC-9 and PC-14 NSCLC cell lines, and cross-resistance to 1 in the cisplatin-resistant cells was largely circumvented. Complex 3 was moderately cytotoxic, whereas 2 and 4 were only marginally cytotoxic. We also determined the growth inhibitory activities of 1 and 3, as well as prototype azolato-bridged complexes [{cis-Pt(NH3)2}2(μ-OH)(μ-pyrazolato)](2+) (AMPZ), [{cis-Pt(NH3)2}2(μ-OH)(μ-1,2,3-triazolato-N1,N2)](2+) (AMTA), [{cis-Pt(NH3)2}2(μ-OH)(μ-tetrazolato-N1,N2)](2+) (5-H-X), and [{cis-Pt(NH3)2}2(μ-OH)(μ-tetrazolato-N2,N3)](2+) (5-H-Y), against a panel of 39 human cancer cell lines (JFCR39). The average 50% growth inhibition concentrations of the complexes against the JFCR39 cell lines ranged from 0.933 to 23.4 μM. The cytotoxicity fingerprints of the complexes based on the JFCR39 cytotoxicity data were similar to one another but completely different from the fingerprints of clinical platinum-based anticancer drugs. Complex 3 exhibited marked antitumor efficiency when tested in vivo on xenografts of PANC-1 pancreatic cancer in nude mice. The high potency of 3 confirmed that the tetrazolato-bridged structure exhibits high in vivo antitumor efficacy.

Characteristic effect of an anticancer dinuclear platinum(II) complex on the higher-order structure of DNA

It is known that a 1,2,3-triazolato-bridged dinuclear platinum(II) complex, [{cis-Pt(NH3)2}2(µ-OH)(µ-1,2,3-ta-N1,N2)](NO3)2 (AMTA), shows high in vitro cytotoxicity against several human tumor cell lines and circumvents cross-resistance to cisplatin. In the present study, we examined a dose- and time-dependent effect of AMTA on the higher-order structure of a large DNA, T4 phage DNA (166 kbp), by adapting single-molecule observation with fluorescence microscopy. It was found that AMTA induces the shrinking of DNA into a compact state with a much higher potency than cisplatin. From a quantitative analysis of the Brownian motion of individual DNA molecules in solution, it became clear that the density of a DNA segment in the compact state is about 2,000 times greater than that in the absence of AMTA. Circular dichroism spectra suggested that AMTA causes a transition from the B to the C form in the secondary structure of DNA, which is characterized by fast and slow processes. Electrophoretic measurements indicated that the binding of AMTA to supercoiled DNA induces unwinding of the double helix. Our results indicate that AMTA acts on DNA through both electrostatic interaction and coordination binding; the former causes a fast change in the secondary structure from the B to the C form, whereas the latter promotes shrinking in the higher-order structure as a relatively slow kinetic process. The shrinking effect of AMTA on DNA is attributable to the possible increase in the number of bridges along a DNA molecule. It is concluded that AMTA interacts with DNA in a manner markedly different from that of cisplatin.

An in vivo highly antitumor-active tetrazolato-bridged dinuclear platinum(II) complex largely circumvents in vitro cisplatin resistance: two linkage isomers yield the same product upon reaction with 9-ethylguanine but exhibit different cytotoxic profiles

Cytotoxicity assays of azolato-bridged dinuclear Pt(II) complexes, [{cis-Pt(NH(3))(2)}(2)(μ-OH)(μ-azolato)](2+), where the azolato was pyrazolato (1), 1,2,3-triazolato-N1,N2 (2), tetrazolato-N1,N2 (3), or tetrazolato-N2,N3 (4), were performed in cisplatin-sensitive and -resistant human non-small-cell lung cancer cell lines (PC-9 and PC-14). These complexes largely circumvented the cisplatin resistance in both cell lines, with resistance factors for 1-4 in the range of 0.5-0.8 and 0.9-2.0 for PC-9 and PC-14 cells, respectively. Complex 4 exhibited approximately 10 times the cytotoxicity of 3. When 3 and 4 were reacted with 2 molar equiv. of 9-ethylguanine (9EtG), they yielded an identical product, [{cis-Pt(NH(3))(2)(9EtG-N7)}(2)(μ-tetrazolato-N1,N3)](3+), that had N1,N3 platinum coordination through a Pt(II) migration process on the tetrazolate ring. The second-order rate kinetics of these isomers were almost the same as each other and faster than those of 1 and 2. The cytotoxicity of azolato-bridged complexes, except for 3, increases as their kinetic rates in the 9EtG reaction increase.

Isolation, characterization and thiol exchange reaction of penicillamine selenotrisulfides

Penicillamine selenotrisulfides formed by the reaction of selenite with D-, L-, and DL-penicillamines in the aqueous solution have been isolated and submitted to the spectroscopic and chromatographic analyses. FAB-MS spectra indicated the molecular ion peak (M+1)+ at m/z 377, which confirmed the molecular formula (C10H20N2O4S2Se) for all these selenotrisulfides. The [77Se]NMR and HPLC results showed that symmetric selenotrisulfides (DD and LL) were formed from D- and L-penicillamines, respectively, while asymmetric selenotrisulfide (DL) in addition to symmetric ones was formed from DL-penicillamine. The occurrence of thiol exchange reaction of selenotrisulfides was verified.