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Recently research shows that horseradish peroxidase, HRP, when combined with other compounds, is highly reactive toward different human tumour cells and that better understanding of catalytic mechanism and inhibition HPR could lead to a new targeted cancer therapy. Thus, the inhibition of HRP activity by dipotassium-trioxohydroxytetrafluorotriborate K2[B3O3F4OH] was investigated for possible explanation of previously observed antitumour activities of this promising drug. HRP activity was studied under steady-state kinetic conditions by a spectrophotometric method. In the absence of the inhibitor values of Km = 0.47 mM and Vmax = 0.34 mM min−1, respectively, were determined. The hydrogen peroxide H2O2 kinetic measurements show a competitive inhibition with the inhibition constant KI = 2.56 mM. The activation energy Ea values were found to be very similar for both reactions; in the absence of inhibitor activation energy was 17.7 kJ mol−1 and in the presence of inhibitor activation energy was 16.3 kJ mol−1. The values of Arrhenius constants were found to be different; A = 4.635 s−1 was measured in the absence of inhibitor while in the presence of inhibitor Arrhenius constant was 1.745 s−1 showing that K2[B3O3F4OH] initiates conformational change in the structure of the HRP and subsequently reduces its activity. [ABSTRACT FROM AUTHOR]

The effect of Ca2+ ions on the cytotoxic ability of boron heterocyclic compound dipotassiumtrioxohydroxytetrafluorotriborate (K2[B3O3F4OH]), on in vitro tumor cells (mammary adenocarcinoma 4T1, melanoma B16F10 and squamous cell carcinoma SCCVII) and non-tumoral fibroblast cells (mouse dermal L929 and hamster lung V79) was examined. At small concentrations of Ca2+ ions (0.42 mM), K2[B3O3F4OH] (3.85 mM) has a very strong cytotoxic effect on all cancer cells tested (89.1, 85.6 and 84.6%) and significantly less effect on normal cells (19.5 and 24.2%), respectively. Applying larger concentrations of Ca2+ ions (9.42 72.42 mM), at the same concentration of K2[B3O3F4OH], no significant cytotoxic effect was detected on cancer cells and normal cells investigated. The selective ability of K2[B3O3F4OH], in the medium with a low concentration of Ca2+ ions has a strong cytotoxic effect on cancer cells and very weak effect in normal cells, opens up the possibility of its application in antitumor therapy. [ABSTRACT FROM AUTHOR]

Recently it was found that dipotassium-trioxohydroxytetrafluorotriborate, K2(B3O3F4OH), is a potent and highly specific inhibitor of precancerous cell processes. We conducted gene expression profiling of human melanoma cells before and after treatment with two concentrations (0.1 and 1 mM) of this boron inorganic derivative in order to assess its effects on deregulation of genes associated with tumor pathways. Parallel trypan blue exclusion assay was performed to assess the cytotoxicity effects of this chemical. Treatment with K2(B3O3F4OH) induced a significant decrease of cell viability in melanoma cellline at both tested concentrations. Furthermore, these treatments caused deregulation of more than 30 genes known as common anti-tumor drug targets. IGF-1 and hTERT were found to be significantly downregulated and this result may imply potential use of K2(B3O3F4OH) as an inhibitor or human telomerase and insulin-like growth factor 1, both of which are associated with various tumor pathways. [ABSTRACT FROM PUBLISHER]

Dipotassium-trioxohydroxytetrafluorotriborate K2[B3O3F4OH] was listed as a promising new therapeutic for cancer diseases. For in vitro and in vivo investigation of its antitumor effects 4T1 mammary adenocarcinoma, B16F10 melanoma and squamous cell carcinoma SCCVII were used. The detailed in vitro investigation undoubtedly showed that K2[B3O3F4OH] affects the growth of cancer cells. The proliferation of cells depends on the concentration so that aqueous solution of K2[B3O3F4OH], the concentrations of 10−4 M and less, does not affect cell growth, but the concentrations of 10−3 M or more, significantly slows cells growth. B16F10 and SCCVII cells show higher sensitivity to the cytotoxic effects of K2[B3O3F4OH] compared to 4T1 cells. Under in vivo conditions, K2[B3O3F4OH] slows the growth of all three tumors tested compared to the control, and the inhibitory effect was most pronounced during the application of the substance. There is almost no difference if K2[B3O3F4OH] was applied intraperitoneally, intratumor, peroral or as ointment. Addition of 5-FU did not further increase the antitumor efficacy of K2[B3O3F4OH]. [ABSTRACT FROM AUTHOR]

The boron heterocyclic compound dipotassium-trioxohydroxytetrafluorotriborate (K2[B3O3F4OH]) was investigated as inhibitor of the zinc enzyme, carbonic anhydrase (CA, EC 4.2.1.1). Eleven human (h) CA isoforms, hCA I-IV, VA, VI, VII, IX and XII-XIV, were included in the investigations. The anion, similar to tetraborate or phenylboronic acid, inhibited most of them. hCA III was not inhibited by K2[B3O3F4OH], whereas hCA VA, hCA VI, hCA IX and hCA XIII were inhibited in the submillimolar range, with KIs of 0.31-0.63 mM. hCA I and II (cytosolic, widespread isoforms), hCA IV (membrane-bound isoform), hCA XII (tumor-associated, transmembrane) and hCA XIV (transmembrane) were much more effectively inhibited by this anion, with inhibition constants ranging from 25 to 93 µM. hCA VII, a cytosolic enzyme present in the brain and associated to oxidative stress, was very effectively inhibited by K2[B3O3F4OH], with a KI of 8.0 µM. We propose that K2[B3O3F4OH] binds to the metal ion from the enzyme active site, coordinating to the Zn(II) ion monodentately through its B-OH functionality. We hypothesize that some of the beneficial antitumor effects reported for K2[B3O3F4OH] may be due to the inhibition of CAs present in skin tumors. [ABSTRACT FROM AUTHOR]

In the development of boronic acid-based enzyme inhibitors as potential pharmaceutical drugs, dipotassium trioxohydroxytetrafluorotriborate K2[B3O3F4OH] was listed as a promising new therapeutic for treatment of these diseases. The catalase-mediated conversion of hydrogen peroxide, in the presence and absence of K2[B3O3F4OH] was studied. The kinetics conformed to the Michaelis-Menten model. Lineweaver-Burk plots were linear and plotted the family of straight lines intersected on the abscissa indicating non-competitive inhibition of the catalase. It appears that in the absence of inhibitor, catalase operates the best at conditions around pH 7.1 and in the presence of K2[B3O3F4OH] the optimum is around pH 6.2. The uncatalyzed reaction of hydrogen peroxide decomposition generally has a value of activation energy of 75 kJ mole−1, whereas catalase, in the absence of inhibitor, lowers the value to 11.2 kJ mole−1, while in the presence 69 mmoles L−1 of K2[B3O3F4OH] it was 37.8 kJ mole−1. [ABSTRACT FROM AUTHOR]

The methanol, ethyl acetate and chloroform extracts of selected Croatian plants were tested for their acetylcholinesterase (AChE) inhibition and antioxidant activity. Assessment of AChE inhibition was carried out using microplate reader at 1 mg mL−1. Antioxidant capacities were determined by 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging test and ferric reducing/antioxidant power assay (FRAP). Total phenol content (TPC) of extracts were determined using Folin–Ciocalteu colorimetric method. Out of 48 extracts, only methanolic extract of the Salix alba L. cortex exerted modest activity towards AChE, reaching 50.80% inhibition at concentration of 1 mg mL−1. All the other samples tested had activity below 20%. The same extract performed the best antioxidative activity using DPPH and FRAP method, too. In essence, among all extracts used in the screening, methanolic extracts showed the best antioxidative activity as well as highest TPC. [ABSTRACT FROM PUBLISHER]

As a part of an investigation of natural antioxidants from Dalmatian aromatic plants, in this paper we report a study of the antioxidant activity related to the chemical composition of savory free volatile compounds. Twenty-one compounds were identified in the essential oil without fractionation, representing 97.4% of the total oil. The major compound was phenolic monoterpene thymol (45.2%). Other important compounds were monoterpenic hydrocarbons p -cymene (6.4%) and γ-terpinene (5.9%) and oxygen-containing compounds carvacrol methyl ether (5.8%), thymol methyl ether (5.1%), carvacrol (5.3%), geraniol (5.0%) and borneol (3.9%). The evaluation of antioxidant power was performed in vitro by the β-carotene bleaching and thiobarbituric acid (TBA) methods. As determined with both methods, the total savory essential oil as well as different fractions or pure constituents containing hydroxyl group exhibited relatively strong antioxidant effect. The hydrocarbons, when isolated as CH fraction, showed the poorest effectiveness in spite the fact that this fraction contained γ-terpinene, α-terpinene, p -cymene and terpinolene which previously were identified as potential antioxidants. [ABSTRACT FROM AUTHOR]

Nereis sarcoplasmic calcium-binding protein contains three functional EF-hand sites which bind Ca2+ or Mg2+ competitively. Here it was confirmed over a large range of [Mg2+] that the positive cooperativity in binding of Ca2+ (nH = 2) is the result of allostery in Mg2+ dissociation. At pH 7.5, Ca2+ or Mg2+ binding provokes the release of 1.4 mol proton/mol protein, whereas no protons are released during Ca2+-Mg2+ exchange. The enthalpy change as a function of Ca2+ binding yields a two-step curve with an inflection point at 1 mol Ca2+/mol protein and a maximum of -66 kJ/mol at 3 mol Ca2+/mol protein. Binding of three Mg2+ ions is cooperative (nH = 1.8) with a maximal enthalpy change of - 15.1 k J/mol protein. Difference spectroscopy led to the conclusion that, in the metal-free protein, the structure around the aromatic residues is well organized, but that Tyr and Trp residues are still solvent-exposed. Upon Ca2+ binding Tyr and Trp spectra are blue-shifted, but some Trp residues are confined to a positively charged pocket. Examination of the Ca2+-saturated threedimensional crystal structure confirmed that Trp4 and Trp57 are located in such pockets or clefts, close to the surface. During the allosteric T → R transition, promoted by binding of the first Mg2+, the Trp residues move to a hydrophobic environment. For both Ca2+ and Mg2+, the enthalpy change and the conformational change in the environment of the aromatic residues is much more pronounced in the first, than in the subsequent two binding steps. In this respect, the latter seem to be equivalent. [ABSTRACT FROM AUTHOR]