U radu se analiziraju proizvodni kapaciteti i proizvodnja sadnog materijala masline u RH, zakonodavni okvir i mjere potpore ovoj proizvodnji te tehnološki aspekt proizvodnje sadnog materijala masline ...u kontekstu izazova pred kojima se nalazi hrvatsko maslinarstvo i maslinarsko rasadničarstvo kao njegov sastavni dio u procesu pristupanja Europskoj uniji. U Hrvatskoj postoji 18 matičnih nasada masline i djeluje 20 registriranih poslovnih subjekata koji proizvode sadnice masline. U matičnim nasadima umatičeno je 3705
stabala od 26 sorti masline. Proizvodnja sadnica masline od 2000. do 2006. godine bilježi neprekidan porast te je u 2006. godini proizvedeno 575.881 sadnica maslina od 22 sorte, a u svim godinama
proizvedeno je najviše sadnica sorte Oblice. U okviru raspoloživih proizvođačkih kapaciteta postoji potencijal za dodatno povećanje domaće proizvodnje. Metoda orošavanja danas je dominantna metoda proizvodnje sadnog materijala masline u svijetu i Hrvatskoj. Metodom orošavanja u Hrvatskoj se proizvodi 90% od ukupnog broja sadnica masline, a domaći proizvođači primjenjuju je u skladu s tehnološkim standardima.
A series of structurally similar 1-substitued heteroaryl fluorene derivatives were prepared in a simple single step reaction, oxidized to fluorenones and then both, fluorenes and fluorenones, were ...methylated to enhance the solubility and increase the affinity to DNA/RNA. Interactions of both, fluorene and fluorenone analogues with various ds-DNA, ds-RNA revealed strong ds-DNA/RNA binding, and various thermal stabilization effects. Most intriguingly, some fluorene derivatives showed opposite fluorescence change (increase for ds-RNA and decrease for ds-DNA), which was not previously reported for any fluorene analogue. CD experiments along with other methods support ds-DNA minor groove binding and major groove ds-RNA binding. All compounds showed negligible interaction with G-quadruplex DNA. Very low cell cytotoxicity of studied compounds combined with very efficient cellular uptake makes these fluorescent dyes safe for laboratory applications. Moreover, especially compounds which show opposite fluorescence response to ds-DNA and ds-RNA, are promising lead compounds for further studies aimed toward ds-RNA-specific fluorescence markers.
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A synthesis of two series of 3-substituted quinazolinones was performed utilizing a green chemistry approach, deep eutectic solvents and microwaves, namely. ...2-Methyl-3-substituted-quinazolin-4(3H)-one derivatives were synthesized in a two-step reaction, using choline chloride:urea deep eutectic solvent (DES). 3-Substituted-quinazolin-4(3H)-ones were synthesized in one-pot one-step reaction of anthranilic acid, amines and orthoester in a microwave reactor. For the synthesis of 2-methyl-3-substituted-quinazolin-4(3H)-ones, first conventional synthesis of benzoxazinone, as an intermediate, was performed. Further, benzoxazinone in reaction with corresponding amines, in choline choline:urea deep eutectic solvent, furnished desired compounds. These procedures are based on green principles with the aim of developing synthetic routes for the potential antitumor agents. All compounds were characterized by LC/MS,
1
H NMR and
13
C NMR spectral techniques. Compound 1 bearing trifluoromethoxyphenyl group showed promising activity against HuT-78 cell line with IC
50
of 51.4 ± 5.1 µM.
Four series of new hybrid molecules with 7-chloroquinoline and arylamidine moieties joined through the rigid -O- (groups I (2a-g) and II (5a-g)) or flexible -NH-CH2-CH2-O- (groups III (8a-g) and IV ...(10a-g)) linker were synthesized, and their DNA/RNA binding properties and cytotoxic activity were tested, against several human cancer lines. The compounds and their interaction with DNA and RNA were studied by UV–Vis and CD spectroscopy. The obtained results showed that the binding affinity of the investigated compounds increases proportionally with the increase of the length and number of groups able to form hydrogen bonds with ds-polynucleotides. Improvement of binding was additionally achieved by reduction of the structural rigidity of the investigated compounds, new hybrid compounds preferentially bind to ctDNA. For most of them the DNA/RNA grooves are dominant binding sites, except for the compounds from group II for which intercalation in polyA-polyU was the dominant binding mode. The antiproliferative effects were tested by the MTT test on normal (MDCK1), carcinoma (HeLa and CaCo2) and leukemia cell lines (Raji and K462). The GI50 values for all investigated compounds ranged from 5 to more than 100 × 10−6 mol dm−3. Carcinoma cells were more resistant to the investigated compounds than leukemia cells. The most effective compounds against leukemia cell lines were from group IV (10a-g), with GI50 values ranging from of 5 and 35 × 10−6 mol dm−3. The cell cycle arrest was investigated by flow cytometry and the obtained results indicate that the selected compounds, 2d, 2e, 8a, 10d, 10e, and 10f, induce changes in the cell cycle of treated cells, but the cycle phase distribution varies between them. A significant decrease in the number of cells in S phase (p < 0.001) was observed in all treated cells, but only 10d and 10f induce cell cycle arrest at G0/G1 phase, dominantly.
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•Four series of novel 7-chloroquinoline-arylamidine hybrids were synthesized.•All compounds bind preferentially to ctDNA.•Antiproliferative activity of the compounds was evaluated in four cancer cell lines.•8a showed strongest inhibitory activity with IG50 = 6.2 μM against Raji cell line.•Compounds 2a,e, 8a, 10d,e,f induced changes in cell cycle of treated cells.
Our recent studies showed significant antiproliferative capacity of newly synthesized N-9-sulfonylpyrimidine derivatives on human carcinoma, lymphoma, and leukemia cells. These derivatives behave as ...antimetabolites exploiting cellular metabolism to induce cytotoxicity in treated tumors’ cells 1-3. Based on previously obtained results on N-9-sulfonylpyrimidine derivatives, a new series of N-9-sulfonylpurine derivatives were synthesized and tested on biological potential.
The aim of this study was to determine if measured antitumor activity of N-9-sulfonylpurine derivatives is linked with ROS accumulation and mitochondrial membrane destabilization in human cervix adenocarcinoma (HeLa), human chronic myelogenous leukemia (K562) and human Burkitt’s lymphoma (Raji) cells. Changes in the mitochondrial membrane potential (∆Ψm) of tested compounds were determined by flow cytometry, measuring fluorescence of JC-1 dye in tumor cells after 24 hours of treatment. Intracellular accumulation of ROS was determined after 1 hour in cells exposed to N-9-sulfonylpurine derivatives by flow cytometry as well. Tested derivatives induced increased accumulation of ROS and mitochondrial disruption in more than 80% of HeLa and Raji cells. The effect of tested derivatives on K562 cells is reflected by the change of mitochondrial potential in more than 70% with slightly reduced accumulation of ROS. Based on obtained results we can conclude that newly synthesized N-9-sulfonylpurine derivatives are good candidates for further antitumor studies.
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•Synthesis of new monomethine, unsymmetrical styryl dyes based on benzothiazole and piperazine scaffolds.•High binding affinities of dyes toward ds-DNA, ds-RNA and G-quadruplexes.•New ...dyes with N-methylpiperazine function showed significant preference for AT-DNA polynucleotide sequence and demonstrated minor groove binding.•New dyes with N-phenylpiperazine function showed high affinities for G-quadruplexes and micromolar and submicromolar antiproliferative activity toward human tumour cell lines.•Accumulation of dye in the mitochondrial space.
New monomethine, unsymmetrical styryl dyes consisting of benzothiazole and N-methylpiperazine or N-phenylpiperazine scaffolds were synthesized, and their binding affinities for different ds-polynucleotides and G-quadruplex were studied. Substitution of piperazine unit with methyl or phenyl group strongly influenced their binding modes, binding affinities, spectroscopic responses and antiproliferative activities. Compounds with N-methylpiperazine substituents showed a significant preference for AT-DNA polynucleotides and demonstrated AT-minor groove binding, which manifested in strong fluorescence increase, significant double helix stabilization, and positive induced circular dichroism spectra. These compounds formed complexes with G-quadruplex by π-π stacking interactions of dye with the top or bottom G-tetrad. Bulkier compounds with N-phenylpiperazine function are probably bound to ds-polynucleotide by partial intercalation between base pairs. On the other hand, they showed stronger stabilization of G-quadruplex compared to methyl-substituted compounds. Fluorimetric titrations pointed to possible mixed stoichiometry’s: 1:1 complex with π-π stacking interactions of dye on the top or bottom G-tetrad and 1:2 complex with dye positioned between two G-quadruplex molecules. Bulkier dyes with N-phenylpiperazine fragments demonstrated micromolar and submicromolar antiproliferative activity that was especially pronounced for leukaemia and lymphoma. Flow cytometric assay shows dose- and time-dependent increase in SubG0/G1 phase. Furthermore, the compounds enter the cells readily and accumulate in the mitochondrial space, co-localize with the standard mitochondrial markers.
In the frame of this paper, the enzyme-assisted hydrolysis coupled with ultrasound and Soxhlet extraction was applied in order to get extracts of chamomile ligulate flowers (CLF). Obtained extracts ...were characterized in terms to their apigenin and apigenin glucoside composition, as well as antiproliferative potential against tumour cells. Antioxidant activity was determined by two different assays based on different mechanisms showing that autofermented extracts have higher reduction potential. Autofermented extracts prepared by ultrasound and Soxhlet extraction had a stronger impact on the treated carcinoma (HeLa and NCI-H358) and leukemia (K562) cells' growth reduction in comparison to the native extracts, 30-35% greater inhibition at the lowest concentration (0.01 mg/mL), in two observed time points (48 and 72 h). Leukemia cells are more sensitive to all tested extracts. The autofermented CLF extracts with highest antiproliferative efficacy induced morphological changes and apoptosis in the HeLa cells. Obtained results clearly showed that the combination of enzymatic hydrolysis with cavitation phenomenon results in extracts with higher apigenin content and increased biological potential.
A series of novel hydrazino-based peptidomimetics and analogues comprising N-terminal lysine and C-terminal phenanthridinyl-l-alanine were prepared. The presented results demonstrate the up to now ...unknown possibility to finely modulate peptide interactions with DNA/RNA by α-hydrazino group insertion and how the different positioning of two α-hydrazino groups in peptides controls binding to various double stranded and single stranded DNA and RNA. All peptidomimetics bind with 1-10 micromolar affinity to ds-DNA/RNA, whereby the binding mode is a combination of electrostatic interactions and hydrophobic interactions within DNA/RNA grooves. Insertion of the α-hydrazino group into the peptide systematically decreased its fluorimetric response to DNA/RNA binding in the order: mono-hydrazino < alternating-hydrazino < sequential-hydrazino group. Binding studies of ss-polynucleotides suggest intercalation of phenanthridine between polynucleotide bases, whereby affinity and fluorimetric response decrease with the number of α-hydrazino groups in the peptide sequence. Particularly interesting was the interaction of two sequential α-hydrazino acids-peptidomimetic with poly rG, characterised by a specific strong increase of CD bands, while all other peptide/ssRNA combinations gave only a CD-band decrease. All mentioned interactions could also be reversibly controlled by adjusting the pH, due to the protonation of the fluorophore.
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In an attempt to enhance the previously observed antiproliferative capacity of 1-(p-toluenesulfonyl)cytosine (N-1-tosylcytosine, ligand 1), its copper(II) complex (Cu(1-TsC-N3)2Cl2, ...complex 2) was prepared and tested in vitro on various carcinoma and leukemia cells. The comparative in vitro studies using the ligand 1, the complex 2, CuCl2x2H2O salt (salt 3) and the 1:2 mixture of the salt 3 and ligand 1 (mixture 4) were performed on normal (WI38), human carcinoma (HeLa, CaCo2, MiaPaCa2, SW620), lymphoma (Raji) and leukemia (K562) cell lines. Significantly elevated concentration of the intracellular copper after treatment of K562 cells and HeLa cells during 2h with complex 2 (7.83 vs. 5.4 times) was detected by atomic absorption spectroscopy. Cytotoxicity was analyzed by MTT assay. We found that antiproliferative capacity of the tested compounds varies (IC50 after 72h of exposure: 0.6×10−6M to>100×10−6M). Leukemia and lymphoma cells were found the most sensitive to complex 2 which showed more than 100 times higher in vitro activity against K562 cells than ligand 1. Apoptotic morphological changes, an externalization of phosphatydilserine, and changes in the mitochondrial membrane potential of treated cells were found. The caspase-3 activity in HeLa and K562 cells was measured by caspase-3 colorimetric assay kit. Caspase-3 was not activated in the treated K562 cells while salt 3 and the mixture 4 in the HeLa cells significantly increased tested enzyme activity. These findings suggest that copper(II) in the molecular complex 2 by improving entry of the N-1-tosylcytosine 1 into cells increases its antiproliferative capacity.
In summary, the present study demonstrated that complex 2 possesses an antileukemic effect on K562 cells, and its anticancer activity was attributed with induction of apoptosis. The exact mechanism of apoptosis induction by complex 2 must be further investigated.
The synthesis of novel 6-chloro/morpholino/amino/-9-sulfonylpurine derivatives was accomplished in two ways, either (i) involving the condensation reaction of 6-chloropurine with commercially ...available arylsulfonyl chlorides in acetone and the presence of aqueous KOH at 0 °C, followed by the substitution of C6-chlorine with morpholine, or (ii) employing a reversed synthetic approach where 6-morpholinopurine and commercially available adenine bases were reacted with the corresponding alkyl, 2-arylethene and arylsulfonyl chlorides giving the N9 sulfonylated products, the latter particularly used where prior nonselective sulfonylation was observed. In both approaches, the sulfonylation reaction occurred regioselectively at the purine N9 position lacking any concurrent N7 derivatives, except in the case of a smaller methyl substituent on SO
2
and the free amino group at C6 of the purine ring. The tautomeric features of initial N9 unsubstituted purines, as well as stability trends among the prepared N-9-sulfonylpurine derivates, were investigated using DFT calculations with an important conclusion that electron-donating C6 substituents are beneficial for the synthesis as they both promote the predominance of the desired N9 tautomers and help to assure the stability of the final products. The newly synthesized 6-morpholino and 6-amino-9-sulfonylpurine derivatives showed antiproliferative activity on human carcinoma, lymphoma, and leukemia cells. Among the tested compounds, 6-morpholino 17 and 6-amino 22 derivatives, with trans-β-styrenesulfonyl group attached at the N9 position of purine, proved to be the most effective antiproliferative agents, causing accumulation of leukemia cells in subG0 cell cycle phase.
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