Synthesis, physicochemical properties and biological activities of novel alkylphosphocholines with foscarnet moiety.

A series of alkylphosphocholines with foscarnet moiety was synthesized. The structure of these zwitterionic amphiphiles was modified in both polar and non-polar parts of surfactant molecule. Investigations of physicochemical properties are represented by the determination of critical micelle concentration, the surface tension value at the cmc and the surface area per surfactant head group utilising surface tension measurements. Hydrodynamic diameter of surfactant micelles was determined using the dynamic light scattering technique. Alkylphosphocholines exhibit significant cytotoxic, anticandidal (Candida albicans) and antiamoebal (Acanthamoeba spp. T4 genotype) activity. The relationship between the structure, physicochemical properties and biological activity of the tested compounds revealed that lipophilicity has a significant influence on biological activity of the investigated surfactants. More lipophilic alkylphosphocholines with octadecyl chains show cytotoxic activity against cancer cells which is higher than that of the compounds with shorter alkyl chains. The opposite situation was observed in case of anticandidal and antiamoebal activity of these surfactants. The most active compounds were found to have pentadecyl chains. The foscarnet analogue of miltefosine C15-PFA-C showed the highest anticandidal activity. The minimum value of anticandidal activity of this compound is 1,4 µM thus representing the highest anticandidal activity found within the group of alkylphosphocholines.

Biological Activities and ADMET-Related Properties of Novel Set of Cinnamanilides.

A series of nineteen novel ring-substituted N-arylcinnamanilides was synthesized and characterized. All investigated compounds were tested against Staphylococcus aureus as the reference strain, two clinical isolates of methicillin-resistant S. aureus (MRSA), and Mycobacterium tuberculosis. (2E)-N-[3-Fluoro-4-(trifluoromethyl)phenyl]-3-phenylprop-2-enamide showed even better activity (minimum inhibitory concentration (MIC) 25.9 and 12.9 µM) against MRSA isolates than the commonly used ampicillin (MIC 45.8 µM). The screening of the cell viability was performed using THP1-Blue™ NF-κB cells and, except for (2E)-N-(4-bromo-3-chlorophenyl)-3-phenylprop-2-enamide (IC50 6.5 µM), none of the discussed compounds showed any significant cytotoxic effect up to 20 µM. Moreover, all compounds were tested for their anti-inflammatory potential; several compounds attenuated the lipopolysaccharide-induced NF-κB activation and were more potent than the parental cinnamic acid. The lipophilicity values were specified experimentally as well. In addition, in silico approximation of the lipophilicity values was performed employing a set of free/commercial clogP estimators, corrected afterwards by the corresponding pKa calculated at physiological pH and subsequently cross-compared with the experimental parameters. The similarity-driven property space evaluation of structural analogs was carried out using the principal component analysis, Tanimoto metrics, and Kohonen mapping.

pH-Sensitive N,N-Dimethylalkane-1-amine N-Oxides in DNA Delivery: From Structure to Transfection Efficiency.

pH-sensitive liposomes composed of homologues of series of N,N-dimethylalkane-1-amine N-oxides (CnNO, n = 8-18, where n is the number of carbon atoms in the alkyl substituent) and neutral phospholipid dioleoylphosphatidylethanolamine (DOPE) were prepared at two molar ratios (CnNO/DOPE = 0.4:1 and 1:1) and tested for their in vitro transfection activity. Several techniques (SAXS/WAXS, UV-vis, zeta potential measurements, confocal microscopy) were applied to characterize the system in an effort to unravel the relationship among the transfection efficiency, structure, and composition of the lipoplexes. The transfection efficiency of CnNO/DOPE for plasmid DNA in U2OS cells follows a quasi-parabolic dependence on CnNO's alkyl substituent length with a maximum at n = 16. The transfection efficiency of CnNO/DOPE (n = 12-18) lipoplexes was found to be higher than that of commercially available Lipofectamine 2000. C16NO/DOPE also positively transfected HEK 293T and HeLa cells. Small-angle X-ray scattering (SAXS) shows large structural diversity depending on the complex's composition and pH. Transfection efficiencies mediated by two structures, either a condensed lamellar (Lαc) or epitaxially connected Lαc and a condensed inverted hexagonal (HIIc) phase (Lαc & HIIc), were found to be very similar. The change in pH from acidic to neutral induces phase transition Lαc & HIIc → QII + Lα, with cubic phase QII of the Pn3m space group. QII detected in lipoplexes of most efficient composition CnNO/DOPE (n = 16 and 18) facilitates DNA release and promotes its internalization in the cell.

Improvement of Glibenclamide Water Solubility by Nanoparticle Preparation.

Glibenclamide, a drug used for the treatment of type 2 diabetes, belongs to Class II of Biopharmaceutical Classification System. It is a highly permeable, but poorly water-soluble drug. Nanoparticles of glibenclamide were prepared by an emulsion solvent evaporation method using dichloromethane as a solvent of glibenclamide and 3% (w/w) aqueous solution of carboxymethyl dextran sodium salt as a stabilizer, which was found as optimal. A solubility test comparing the water solubility of glibenclamide bulk and nanoparticles confirmed the improved (2-fold higher) solubility of glibenclamide nanoparticles (0.045 µg/ml) compared with bulk (0.024 µg/ml).

Self-Assembly Properties of Cationic Gemini Surfactants with Biodegradable Groups in the Spacer.

: Self-assembly properties of cationic gemini surfactants with biodegradable amide or ester groups in the spacer were investigated utilising time-resolved fluorescence quenching, dynamic light scattering and zeta potential measurements. A correlation between aggregation parameters such as micelle aggregation number, micelle size and zeta potential with the structure of gemini molecules was made. For gemini molecules with medium spacer lengths, micelle aggregation number does not change much with the surfactant concentration. When the spacer is extended, a stronger aggregation tendency is observed for gemini surfactant molecules with two ester groups in the spacer and the aggregation number increases. The assumption of stronger aggregation of ester-based gemini molecules at larger spacer number values is also documented by measurements of the size and zeta potential of ester-based micelles. The explanation of the difference in aggregation ability of amide-based and ester-based gemini molecules is related to the structural features of gemini molecules, notably to the larger flexibility and denser arrangement of ester-based gemini molecules in a micelle. To support this assumption, optimised 3D models of the studied gemini molecules were constructed. Correspondingly, the calculations show smaller size and interfacial area for ester-based gemini conformers.

The Synthesis, Self-Assembled Structures, and Microbicidal Activity of Cationic Gemini Surfactants with Branched Tridecyl Chains.

Cationic gemini surfactants with polymethylene spacer and linear alkyl chains containing an even number of carbon atoms have been extensively studied in the recent past, with the emphasis put on the determination of their aggregation behaviour in aqueous solution and their biological properties. However, the information on the aggregation of branched gemini surfactants with an odd number of carbon atoms in their alkyl chains is only sparsely reported in the literature. To help cover this gap in the research of cationic gemini surfactants, a series of branched bisammonium cationic gemini surfactants with an odd number of carbon atoms in alkyl chains (tridecane-2-yl chains) and a polymethylene spacer with a variable length ranging from 3 to 12 carbon atoms have been synthesized and investigated. Critical micelle concentration, which was determined by three methods, was found to be in the order 10-4 mol/L. A comparison of the obtained data of the novel series of tridecyl chain geminis with those of gemini surfactants with dodecyl chains and an identical spacer structure revealed that structural differences between both series of gemini surfactants result in different aggregation and surface properties for surfactants with 6 and 8 methylene groups in the spacer (N,N'-bis(tridecane-2-yl)-N,N,N',N'-tetramethylhexane-1,6-diaminium dibromide and N,N'-bis(tridecane-2-yl)-N,N,N',N'-tetramethyloctane-1,8-diaminium dibromide) with the cmc values 8.2 × 10-4 mol/L and 6.5 × 10-4 mol/L, respectively, as determined by surface tension measurements. Particle size analysis showed the formation of small stable spherical micelles in the interval between 2.8 and 5 nm and with zeta potential around +50 mV, which are independent of surfactant concentration and increase with the increasing spacer length. Microbicidal activity of 13-s-13 gemini surfactants was found to be efficient against Gram-positive, Gram-negative bacteria and yeast.

Investigation of Anti-Inflammatory Potential of N-Arylcinnamamide Derivatives.

A series of sixteen ring-substituted N-arylcinnamanilides, previously described as highly antimicrobially effective against a wide spectrum of bacteria and fungi, together with two new derivatives from this group were prepared and characterized. Moreover, the molecular structure of (2E)-N-(2-bromo-5-fluorophenyl)-3-phenylprop-2-enamide as a model compound was determined using single-crystal X-ray analysis. All the compounds were tested for their anti-inflammatory potential, and most tested compounds significantly attenuated the lipopolysaccharide-induced NF-κB activation and were more potent than the parental cinnamic acid. (2E)-N-[2-Chloro-5-(trifluoromethyl)phenyl]-3-phenylprop-2-enamide, (2E)-N-(2,6-dibromophenyl)- 3-phenylprop-2-enamide, and (2E)-N-(2,5-dichlorophenyl)-3-phenylprop-2-enamide demonstrated the highest inhibition effect on transcription factor NF-κB at the concentration of 2 µM and showed a similar effectiveness as the reference drug prednisone. Several compounds also decreased the level of TNF-α. Nevertheless, subsequent tests showed that the investigated compounds affect neither IκBα level nor MAPKs activity, which suggests that the N-arylcinnamanilides may have a different mode of action to prednisone. The modification of the C(2,5)' or C(2,6)' positions of the anilide core by rather lipophilic and bulky moieties seems to be preferable for the anti-inflammatory potential of these compounds.

DNA-DOPE-gemini surfactants complexes at low surface charge density: from structure to transfection efficiency.

DNA condensation, structure and transfection efficiency of complexes formed by gemini surfactants alkane-α,ω-diyl-bis(dodecyldimethylammonium bromide)s (CnGS12, n = 3, 6 and 12 is the number of alkane spacer carbons), dioleoylphosphatidylethanolamine (CnGS12/DOPE = 0.3 mol/mol) and DNA at low surface charge density were investigated through different techniques. Small angle X-ray diffraction showed a condensed lamellar phase with marked dependence of DNA-DNA distance on (+/-) charge ratio. High ionic strength of hydrating medium screens the interaction DNA - CnGS12/DOPE and complexed DNA represented maximally ~ 45-60% of total DNA in the solution as derived from fluorescence and UV-VIS spectroscopy. The in vitro transfection efficiency of CnGS12/DOPE liposomes on mammalian HEK 293 cell line was spacer length-dependent. C12GS12/DOPE/DNA complexes exhibited the best transfection efficiency (~ 18% GFP-expressing cells relative to all viable cells) accompanied by ~ 89% cell viability.

Synthesis, structural characterization and biological activity of novel Knoevenagel condensates on DLD-1 human colon carcinoma.

Biologically active Knoevenagel condensates (1-14) of diarylheptanoids: 1,7-bis(3-methoxy-4-hydroxyphenyl)hepta-1,7-diene-3,5-dione and 1,7-bis(3-ethoxy-4-hydroxyphenyl)hepta-1,7-diene-3,5-dione, were synthesized and structurally characterized. Compounds 1-14 exhibited cytotoxicity against colon carcinoma cells, and their antiproliferative effect was associated with a significant decrease of multidrug resistance proteins. One of the underlying mechanisms of these effects is the reduction of intracellular and extracellular SOD enzymes by compounds 1, 12 and 14, which render the tumor cells more vulnerable to oxidative stress.

Influence of quercetin on the interaction of gliclazide with human serum albumin - spectroscopic and docking approaches.

Protein-binding interactions are displacement reactions which have been implicated as the causative mechanisms in many drug-drug interactions. Thus, the aim of presented study was to analyse human serum albumin-binding displacement interaction between two ligands, hypoglycaemic drug gliclazide and widely distributed plant flavonoid quercetin. Fluorescence analysis was used in order to investigate the effect of substances on intrinsic fluorescence of human serum albumin (HSA) and to define binding and quenching properties of ligand-albumin complexes in binary and ternary systems, respectively. Both ligands showed the ability to bind to HSA, although to a different extent. The displacement effect of one ligand from HSA by the other one has been described on the basis of the quenching curves and binding constants comparison for the binary and ternary systems. According to the fluorescence data analysis, gliclazide presents a substance with a lower binding capacity towards HSA compared with quercetin. Results also showed that the presence of quercetin hindered the interaction between HSA and gliclazide, as the binding constant for gliclazide in the ternary system was remarkably lower compared with the binary system. This finding indicates a possibility for an increase in the non-bound fraction of gliclazide which can lead to its more significant hypoglycaemic effect. Additionally, secondary and tertiary structure conformational alterations of HSA upon binding of both ligands were investigated using synchronous fluorescence, circular dichroism and FT-IR. Experimental data were complemented with molecular docking studies. Obtained results provide beneficial information about possible interference upon simultaneous co-administration of the food/dietary supplement and drug.

Silver Nanoparticles Stabilised by Cationic Gemini Surfactants with Variable Spacer Length.

The present study is focused on the synthesis and investigation of the physicochemical and biological properties of silver nanoparticles stabilized with a series of cationic gemini surfactants having a polymethylene spacer of variable length. UV-VIS spectroscopy, dynamic light scattering, scanning electron microscopy and zeta potential measurements were applied to provide physicochemical characterization of the silver nanoparticles. The mean size values of the nanoparticles were found to be in the 50 to 115 nm range. From the nanoparticle size distributions and scanning electron microscopy images it results that a population of small nanoparticles with the size of several nanometers was confirmed if the nanoparticles were stabilized with gemini molecules with either a short methylene spacer (two or four -CH2- groups) or a long spacer (12 -CH2- groups). The average zeta potential value for silver nanoparticles stabilized with gemini molecules is roughly independent of gemini surfactant spacer length and is approx. +58 mV. An interaction model between silver nanoparticles and gemini molecules which reflects the gained experimental data, is suggested. Microbicidal activity determinations revealed that the silver nanoparticles stabilized with gemini surfactants are more efficient against Gram-negative bacteria and yeasts, which has a direct relation to the interaction mechanism of nanoparticles with the bacterial cell membrane and its structural composition.

Novel Palladium(II) Complexes that Influence Prominin-1/CD133 Expression and Stem Cell Factor Release in Tumor Cells.

New Pd(II) complexes of 1,7-bis(2-methoxyphenyl)hepta-1,6-diene-3,5-dione were synthesized and structurally characterized. The complexes were tested in vitro on human colon and hepatic carcinoma cell lines, normal hepatic cells and hematopoietic progenitor cells. Biological tests proved that Pd(II) complexes 1 and 2 (containing a curcumin derivative) exhibit a strong in vitro antitumor effect against the cells derived from human colorectal carcinoma and the hepatic metastasis of a colorectal carcinoma. Complex 1 has an outstanding inhibitory effect against BRAF-mutant colon carcinoma and hepatocarcinoma cell growth; 1 and 2 are both more active than the free ligand and have the capacity to trigger early apoptotic processes. By flow cytometric measurements, an important decrease of prominin-1 (CD133) molecule expression on tumor cells membrane was identified in cell populations subjected to 1 and 2. Quantitative immune enzymatic assay proved restrictions in stem cell factor (SCF) release by treated tumor cells. Although less cytotoxic, the free ligand inhibits the surface marker CD133 expression in hepatocarcinoma cells, and in HT-29 colon carcinoma. The new synthesized Pd(II) complexes 1 and 2 exhibit an important potential through their selective cytotoxic activity and by targeting the stem-like tumor cell populations, which leads to the tumor growth arrest and prevention of metastasis.

Stimuli responsive polymorphism of C12NO/DOPE/DNA complexes: Effect of pH, temperature and composition.

N,N-dimethyldodecylamine-N-oxide (C12NO) is a surfactant that may exist either in a neutral or cationic protonated form depending on the pH of aqueous solutions. Using small angle X-ray diffraction (SAXD) we observe the rich structural polymorphism of pH responsive complexes prepared due to DNA interaction with C12NO/dioleoylphosphatidylethanolamine (DOPE) vesicles and discuss it in view of utilizing the surfactant for the gene delivery vector of a pH sensitive system. In neutral solutions, the DNA uptake is low, and a lamellar Lα phase formed by C12NO/DOPE is prevailing in the complexes at 0.2≤C12NO/DOPE<0.6 mol/mol. A maximum of ~30% of the total DNA volume in the sample is bound in a condensed lamellar phase LαC at C12NO/DOPE=1 mol/mol and pH7.2. In acidic conditions, a condensed inverted hexagonal phase HIIC was observed at C12NO/DOPE=0.2 mol/mol. Commensurate lattice parameters, aHC≈dLC, were detected at 0.3≤C12NO/DOPE≤0.4 mol/mol and pH=4.9-6.4 suggesting that LαC and HIIC phases were epitaxially related. While at the same composition but pH~7, the mixture forms a cubic phase (Pn3m) when the complexes were heated to 80°C and cooled down to 20°C. Finally, a large portion of the surfactant (C12NO/DOPE>0.5) stabilizes the LαC phase in C12NO/DOPE/DNA complexes and the distance between DNA strands (dDNA) is modulated by the pH value. Both the composition and pH affect the DNA binding in the complexes reaching up to ~95% of the DNA total amount at acidic conditions.

Preparation of Hydrochlorothiazide Nanoparticles for Solubility Enhancement.

Nanoparticles can be considered as a useful tool for improving properties of poorly soluble active ingredients. Hydrochlorothiazide (Class IV of the Biopharmaceutical Classification System) was chosen as a model compound. Antisolvent precipitation-solvent evaporation and emulsion solvent evaporation methods were used for preparation of 18 samples containing hydrochlorothiazide nanoparticles. Water solutions of surfactants sodium dodecyl sulfate, Tween 80 and carboxymethyl dextran were used in mass concentrations of 1%, 3% and 5%. Acetone and dichloromethane were used as solvents of the model compound. The particle size of the prepared samples was measured by dynamic light scattering. The selected sample of hydrochlorothiazide nanoparticles stabilized with carboxymethyl dextran sodium salt with particle size 2.6 nm was characterized additionally by Fourier transform mid-infrared spectroscopy and scanning electron microscopy. It was found that the solubility of this sample was 6.5-fold higher than that of bulk hydrochlorothiazide.

Antimicrobial Activity and Urease Inhibition of Schiff Bases Derived from Isoniazid and Fluorinated Benzaldehydes and of Their Copper(II) Complexes.

In order to evaluate the influence of substitution on biological properties of Schiff bases and their metal complexes, a series of differently substituted fluorine-containing Schiff bases starting from the drug isoniazid (isonicotinylhydrazide) were prepared and their structures were established by single-crystal X-ray diffraction. Also, four copper(II) complexes of these Schiff bases were synthesized. The prepared compounds were evaluated for their antimicrobial activity and urease inhibition. Two of the Schiff bases exerted activity against C. albicans. All copper(II) complexes showed excellent inhibitory properties against jack bean urease, considerably better than that of the standard inhibitor acetohydroxamic acid.

Cholesterol protects phosphatidylcholine liposomes from N,N-dimethyl-1-dodekanamine N-oxide influence.

The interaction of N,N-dimethyl-1-dodekanamine N-oxide (C12(CH3)2NO) with egg yolk phosphatidylcholine (EYPC) liposomes containing cholesterol (CHOL) was studied. The perturbation of CHOL-EYPC bilayers in unilamellar liposomes (ULL) was observed by the leakage of fluorescent probe calcein. Weak leakage is observed at low surfactant concentration cC12NO (minimal perturbation of the bilayer) followed by an intensive leakage at a middle cC12NO (creation of pores). No change in fluorescence intensity was measured at high cC12NO (calcein totally released from liposomes). The higher CHOL amount in the bilayer, the more surfactant is needed to create pores in the bilayer. Solubiliazation of CHOL-EYPC ULL induced by C12NO was studied turbidimetrically. The solubilization curve consists of three parts: saturation of bilayer at low cC12NO (liposomes are preserved), followed by solubilization (liposome - mixed micelle transition) and post-solubilization. The c12NO concentration needed for the onset of the soubilization raises with the increase of nCHOL:nEYPC. The structure of liposomes is still preserved at total calcein release for all nCHOL:nEYPC.

Effects of N,N-dimethyl-N-alkylamine-N-oxides on DOPC bilayers in unilamellar vesicles: small-angle neutron scattering study.

Small-angle neutron scattering data were collected from aqueous dispersions of unilamellar vesicles (ULVs) consisting of mixtures of 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine and a homologous series of N,N-dimethyl-N-alkylamine-N-oxides (CnNO, n = 12, 14, 16, and 18, where n is the number of carbon atoms in the alkyl chain). A modeling approach was applied to the neutron scattering curves to obtain the bilayer structural parameters. Particularly, the external (2)H2O/H2O contrast variation technique was carried out on pure dioleoylphosphatidylcholine (DOPC) ULVs to determine the hydrophilic region thickness [Formula: see text] = 9.8 ± 0.6 Å. Consequently, the hydrocarbon region thickness [Formula: see text], the lateral bilayer area per one lipid molecule [Formula: see text], and the number of water molecules located in the hydrophilic region per one lipid molecule [Formula: see text] were obtained from single-contrast neutron scattering curves using the previously determined [Formula: see text]. The structural parameters were extracted as functions of [Formula: see text] (the CnNO:DOPC molar ratio) and n. The dependences [Formula: see text] provided the partial lateral areas of CnNOs ([Formula: see text]) and DOPC ([Formula: see text]) in bilayers. It was observed that the [Formula: see text]'s were constant in the investigated interval of [Formula: see text] and for n = 12, 14, and 16 equal to 36.6 ± 0.4 Å(2), while [Formula: see text] increased to 39.4 ± 0.4 Å(2). The bilayer hydrocarbon region thickness [Formula: see text] decreased with intercalation of each CnNO. This effect increased with [Formula: see text] and decreased with increasing CnNO alkyl chain length. The intercalation of C18NO changed the [Formula: see text] only slightly. To quantify the effect of CnNO intercalation into DOPC bilayers we fit the [Formula: see text] dependences with weighted linear approximations and acquired their slopes [Formula: see text].

Preparation of risedronate nanoparticles by solvent evaporation technique.

One approach for the enhancement of oral drug bioavailability is the technique of nanoparticle preparation. Risedronate sodium (Biopharmaceutical Classification System Class III) was chosen as a model compound with high water solubility and low intestinal permeability. Eighteen samples of risedronate sodium were prepared by the solvent evaporation technique with sodium dodecyl sulfate, polysorbate, macrogol, sodium carboxymethyl cellulose and sodium carboxymethyl dextran as nanoparticle stabilizers applied in three concentrations. The prepared samples were characterized by dynamic light scattering and scanning electron microscopy. Fourier transform mid-infrared spectroscopy was used for verification of the composition of the samples. The particle size of sixteen samples was less than 200 nm. Polysorbate, sodium carboxymethyl dextran and macrogol were determined as the most favourable excipients; the particle size of the samples of risedronate with these excipients ranged from 2.8 to 10.5 nm.

The DNA-DNA spacing in gemini surfactants-DOPE-DNA complexes.

Gemini surfactants from the homologous series of alkane-α,ω-diyl-bis(dodecyldimethylammonium bromide) (CnCS12, number of spacer carbons n=2-12) and dioleoylphosphatidylethanolamine (DOPE) were used for cationic liposome (CL) preparation. CLs condense highly polymerized DNA creating complexes. Small-angle X-ray diffraction identified them as condensed lamellar phase L(α)(C) in the studied range of molar ratios CnGS12/DOPE in the temperature range 20-60°C. The DNA-DNA distance (d(DNA)) is studied in dependence to CnGS12 spacer length and membrane surface charge density. The high membrane surface charge densities (CnGS12/DOPE=0.35 and 0.4mol/mol) lead to the linear dependence of d(DNA) vs. n correlating with the interfacial area of the CnGS12 molecule.

The microstructure of DNA-egg yolk phosphatidylcholine-gemini surfactants complexes: effect of the spacer length.

BACKGROUND: The length of spacer of gemini surfactants affects the DNA packing in DNA-neutral phospholipid-gemini surfactant complexes. METHODS: The microstructure of complexes DNA-egg yolk phosphatidylcholine (EYPC)-alkane-α,ω-diyl-bis(dodecyl\xaddimethylammonium bromides) (CnGS, spacer n=2-12, n is even) was studied using small angle X-ray diffraction. RESULTS: At EYPC:CnGS=1:1 mol/mol, the condensed lamellar phase was identified in complexes with CnGS, n=2-4, whereas longer spacer (n≥6) induced a hexagonal phase. The condensed lamellar phase Lαc was observed in the range 2\≤ EYPC:GnGS≤10 (mol/mol) in all complexes. The distance between adjacent DNA strands increases linearly with decreasing surface charge density of EYPC-CnGS vesicles. We determined the increase in dDNA 0.40±0.03 nm/1 mol of EYPC from the slope of dDNA=f (molEYPC/molCnGS) in the range of molar ratios 2≤EYPC:CnGS≤5. At lower surface charge density, EYPC:CnGS>5 mol/mol, the length of CnGS spacer (n=6-10) modulates the DNA-DNA distance. CONCLUSIONS: Both the short spacer of CnGS and the low molar ratio EYPC:CnGS result in the closest DNA-DNA packing. A high surface charge density of membrane was reported as a key parameter for transfection efficiency of Lαc phase-forming complexes.