INTRODUCTION:

The innovation in the field of creating various pharmacological and biological activities in compounds that improve the patient’s quality of life and have to offer minimum-reported side effects on patients, represents the main concern for synthetic organic chemists.¹

One of such compounds are acetophenones and their derivatives that are useful compound in chemistry, development of drug, medicine, etc. Their diverse functional groups allow them to display specific biological activities like anticancer, antitubercular, antimicrobial, antifungal, antibacterial, anti-inflammatory, antiviral, and anti-hyperglycemic. These characteristics make them useful in various studies as well as in therapeutic applications.² Acetophenones derivatives are an attractive model molecule when used as foreign substrates for biotransformation, since a readily detectable enantiomer can be formed, and easily determined. These substances have been successfully applied in the building blocks for pharmacological asymmetric synthesis.³ 2’-Hydroxyacetophenone are thought to be crucial intermediates in the synthesis of numerous biological active compounds, especially in the formation of benzopyran compounds, for example the compounds of flavonoids, coumarins derivatives or chromones, these being privileged structures for searching new drug.⁴ Williamson ether synthesis has been widely used in pharmaceutical research, such as the synthesis of anti-influenza drugs, because it involves bimolecular nucleophilic substitution on saturated carbons.⁵ It include the coupling between alkoxides and alkyl halides through a SN2 reaction under basic conditions. Developed by the chemist Alexander Williamson over 160 years ago.⁶ It is considered as one of the reactions classes most often used in synthetic organic chemistry, and is still the best versatile method for the synthesis of unsymmetrical as well as symmetrical ethers.⁷ Alcohol-functional dialdehydes have been previously synthesized via the Williamson ether synthesis, they are used as predecessor for the synthesis of macrocyclic ligand ported a pendent arm, because they may frequentely be induced to undergo Schiff base cyclization reactions when they react with diamines. These pendent arms macrocycles ligand have attracted considerable attention in bioinorganic chemistry as models for biomolecules, and in model chemical science such as in magnetic resonance imaging.⁸

In the present study, Williamson ether reaction of 2’hydroxyacetophenone with 1,3-dichloropropanol in 1:2 mole ratio was used to generate the required bis-ketone 1,3-Bis(2-acetylphenoxy)-2-propanol (denoted as Bis-AcPh) containing two acetophenoxy groups, and ported hydroxyl pendant arm. The structure of the synthesized bis-ketone was determined by FTIR, UV-Vis, and NMR spectroscopic methods. Their biological properties were evaluated by cytotoxicity, antioxidant and anti-inflammatory activities. In Addition, the compound was subjected to chemical reactivity prediction using a DFT study.

MATERIALS AND METHODS:

Experimental animals: In this experience, the procedures for all animal handling followed the guidelines of the Animal Welfare Act, and were performed as stated the guide for the care and the use of laboratory animals of the" Institut Pasteur d’Algérie." Healthy whites mice (female mice, 25-30g weight) were kept in clean cages for fifteen days with acclimatization, and under controlled conditions of 12h light: 12h dark cycle and 24±one°C with free access to standard rodent diet, and clean water before doing the tests.

Synthesis:

The compound 1,3-bis(2-acetylphenoxy)-2-propanol (Bis-AcPh) was prepared according to the method of Lindoy and Armstrong⁹, which was used to synthesize a slightly modified dialdehyde. An ethanolic solution of 2′-hydroxyacetophenone (6.73g, 0.05mol) was added to a 100ml of stirring aqueous sodium hydroxide solution (2.0g, 0.05mol). The mixture was warmed, and then an ethanolic solution of 1, 3-dichloro-2-propanol (3.16g, 0.025mol) was added. In this mixture, enough ethanol (50ml ) was then added to achieve a homogenous solution. The solution was refluxed for 72h, then cooled at 0°C. The bright solid formed was recrystallized using ethanol-water mixture and the resulting brown crystals were filtered and then dried in a vacuum, (yield 44%, melting point: 103°C).

Computational studies:

Basing on the density functional theory (DFT)¹⁰, the Gaussian 09 software performs the theoretical calculations of the optimization of the Bis-AcPh molecular structures, the mapping electrostatic potential, and the frontier molecular orbitals (FMOs). The same theoretical parameters reported in previous papers were used for all calculations.^11,12,13 From the FMOs’ energies, some mathematic characteristic global quantum chemical descriptors (GQDs) were computed according to the formulas (1) to (8).¹⁴

Energy gap (E_GAP)	DE = E_LUMO - E_HOMO	(1)
Ionisation potential (I)	-E_HOMO	(2)
Electron affinity (A)	-E_LUMO	(3)
Electronegativity (c)	χ =-[1/2(E_LUMO+ E_HOMO)]	(4)
Chemical potential (µ)	µ = [1/2(E_LUMO+ E_HOMO)]	(5)
Global hardness (h)	h = 1/2(E_LUMO- E_HOMO)	(6)
Electrophilicity (w)	ω = χ²/2η	(7)
Global softness (s)	σ = 1/ η	(8)

In vitro antioxidant assays: In the DPPH radical scavenging assay, 250μL of the compound Bis-AcPh prepared in methanol at various concentration was added to an equal volume of a methanolic solution of a freshly prepared DPPH (0.004% w/v). The resulting composition was incubated in darkness for a half-hour at room temperature, and Butylated hydroxyl toluene (BHT) was employed as standard. The test was executed in triplicates, and the absorbance was mesured at 517nm against a blank with the aid of spectrophotometer. The decrease in absorbance indicates an increased in radical scavenging activity.¹⁵ Furthermore, the reducing power test is depended on the reduction of Fe^(III) to Fe^(II) according to the following protocol: a portion of 400μL of the compound Bis-AcPh with various concentration was reacted with an equal volume of both phosphate buffer (0.2M, pH = 6.6) and potassium ferricyanide (1%) solutions. The resulting mixture was left to incubate at 50°C for 20min in a water bath. After this time, this reaction was halted by adding a volume of 400μL of the reactif trichloroacetic acid (10%), and then subsequently centrifuging this mixture at 3000rpm for 10min. Finally, 400μL of distilled water, and 80μL of ferric chloride solution (0.1%), where added to 400μL the upper layer of this mixture. After 10min of incubation, the absorbance of this mixture was detected at 700nm, the ascorbic acid (VitC) was considered as reference drug. Higher reaction mixture’s absorbance indicate a better reducing power.¹⁶

Anti-inflammatory activity:

In vitro anti-inflammatory assay by egg albumin denaturation:

The egg albumin denaturation technique was performed by following the process outlined by Karbab et al.¹⁷ with slight modifications. A solution of (1%) of fresh hen’s egg albumin prepared in Tris-HCl (20mM, pH 6.87) buffer solution (v/v) was stirred for 10 min, and then filtered through a strip agase. A volume of 500μL of this prepared albumin solution was mixed with different concentrations of 250μL of the Bis-AcPh or 250μL of Aspirin the reference drug, and the buffer Tris-HCl served as a negative control. These mixtures were kept for incubation at 37°C for 15 min, heated afterwards to 70°C for 5min. The absorbance was then read at 650nm.¹⁷

Topical in vivo anti-inflammatory activity:

In the xylene-induced ear edema assay, animals were assigned randomly to three groups; each group comprised six animals as follows: Group1 (Positive control) was treated topically by indomethacin as the standard drug at a dose of 2mg/ear); group 2 (Negative control) treated topically by xylene (30μL/ear) and group 3 received the same dose (2mg/ear) of Bis-AcPh that was applied immediately after xylene application. All treatment where applied to both the inner and the outer surface of the right ear lobe, the left ear was served as control, and the ear thickness was measured before and after two hours of the last dose of treatment by using a digital calliper.¹⁸ Following the same animal design of topical inflammation induced by xylene; in the croton oil-induced ear edema assay, mice were randomly allocated to three groups of six mice each. About 80μg of croton oil prepared in 15μL of a mixture of acetone/water (1:1, v/v) was applied topically on both the inner and the outer part of each mouse’s right ear at first. Simultaneously, 15μL of an ethanol-water mixture containing 2mg of Bis-AcPh or 2mg of indomethacin, was tested topically at the same region of the ear edema in each mouse, while the negative control group was provoked topically by croton oil. The ear’s thickness was determined in micrometer(µm) using a digital calliper, positioned near the ear’s tip just distal from the cartilaginous ridges. The variation in thickness of mice ear edema before application of the treatment, and after 6 hours of inflammation induced by croton oil treatment was calculated.¹⁹

In vitro cytotoxicity assay:

The method used to evaluate the toxicity of the Bis-AcPh was based on the ability of RBCs to release haemoglobin. Human erythrocytes are prepared from the peripheral blood of a healthy donor in accordance with the standard operating procedures of the International Federation of Blood Donor Organisations (IFBDO). The blood collected in a heparinised tube was subjected to centrifugation at 3000rpm for 10min. The erythrocyte suspension was obtained by washing the cells in 0.9% sterile saline solution, following each washing steps, the pellets containing the cells were spun down at 300rpm for 5min for removing the supernatant. This operation was done three to four times until the supernatant achieved a colourless state. The resultant pallet obtained after the last centrifugation, was quantified and reconstituted using an isotonic buffer solution (10mM sodium phosphate buffer pH 7.4) to prepare a 2%(v/v) suspension. The Bis-AcPh (250µL) prepared at various concentrations, was added to the same volume of prepared erythrocyte suspension .The contents were then incubated for 1h at 37°C, then centrifuged for 10min. The absorbance of the supernatant was read at 540nm against a reagent blank in which the Bis-AcPh was replaced by saline solution.²⁰

Statistical analysis:

Statistical analysis was performed using GraphPad Prism software (version 5.03 for Windows). A one-way analysis of variance (ANOVA) was applied to analyze the data, with results presented as mean±standard deviation (SD). IC₅₀ values were determined through linear regression, and statistical significance was assessed with a confidence level of p<0.05. In vivo experimental results are presented as mean±standard error of the mean (SEM).

RESULTS AND DISCUSSION:

Synthesis and characterization: As described in Scheme 1, the condensation of 2′-hydroxyacetophenone with 1,3-dichloro-2-propanol in a 2:1 M ratio according to Williamson ether reaction afforded the bis-ketone 1,3-Bis (2-acetylphenoxy)-2-propanol (Bis-AcPh).

Scheme 1: Synthesis of Bis-AcPh.

In order to ascertain the structure of this compound, the spectrum of Bis-AcPh demonstrate the typical absorption bands of the functional groups existing in the synthesized product as can be seen from Figure 1. The emergence of the strong vibration bond of (OH) at 3460 cm^-1, together with (C=O) at 1660 cm^-1 and the two asymmetric and symmetric intense bands characteristic of (Ar-O-CH₂) group located at around 1240, 1047 cm^-1respectively, prove the formation of the organic compound Bis-AcPh. The presence of a series of weak bands at 2939, 2876, 2835 cm^-1are due to the vibrations of aliphatic methylene - (CH₂)₃, whereas the intense band located at 1596 cm^-1 represent the stretching vibration (C=C) of the aromatic ring.

Figure 1: The IR spectrum of Bis-AcPh.

Electronic spectrum (Figure 2) of the compound Bis-AcPh recorded in ethanol shows three bands; that centred at 212 nm is ascribed to π-π* electronic transitions of the (C=C), and the (C-C) bonds within the aromatic ring. The band situated at 248 nm is attributed to the n-π* transition of the C=O bond, while the third bond located at 293nm represent the n-σ*of the simple bond C-O.

Figure 2: Electronic Spectrum of Bis-AcPh.

Density functional theory:

Molecular geometry: The Figure 3.a. shows the 3D-molecular geometry structure of the Bis-AcPh that was perfected theoretically by employing DFT. The molecule is deformed from the planar configuration; the angle between the support planes of the aromatic rings has the value 46.48° (Figure 3.b). Furthermore, the lengths of C-C and C=C bonds are in the normal ranges and around 120° indicating sp2 hybridization for all C atoms, and the low difference can be explained by the VSEPR theory.²¹ The results of the DFT calculations agree well with resembling previous molecular structures.^22,23

Figure 3: (a) DFT-Optimized molecular structures: BZT organic part, (b) distortion of the molecular structure.

Mapping electrostatic potential:

The electrostatic potential mapped with the total electron density for the Bis-AcPh molecule is shown in the Figure 4, the surface reveals a positive potential around hydrogen and alkyl carbon, whereas the most negative potential highlighted by a red color is located around the central oxygen atoms of the molecule. This last site can attract nucleophilic compounds. Moreover, a weak negative potential surrounding the cycle revealed by a yellow color can be observed and is related to the π-conjugated electrons. The non-homogeneity of the potential distributions generates a dipolar moment of 1.583Debye.

Figure 4: The total electron density mapped with the electrostatic potential of the Bis-AcPh.

Frontier molecular orbitals (FMOs):

FMOs (HOMO: Highest Occupied Molecular and LUMO: Lowest Unoccupied Molecular Orbitals) are terminology related to the electron transfer phenomena within the surrounding medium, since the HOMO can give electrons, and the LUMO has the preference to receive them.²⁴ The figure 5. displays the plotted HOMO and LUMO of the titled compound. It is seen that, the LUMO is located on the aromatic cycle and its surrounding, while the HOMO is delocalized throughout the molecule except the methyl groups. These results show that the most actives sites for transferring electrons to the surrounding medium are essentially the aromatic cycle. In addition, for the purpose of estimating the electrons transfer; the reactivity and the stability of the Bis-AcPh, the energies of LUMO and HOMO and some related Global quantum physic-chemical descriptors (GQCDs) are an important tool (Table 1). The LUMO and HOMO energies are 5.971Ev and-1.135 respectively (Table 1), the low value of the HOMO’s energy indicates the electron donating character.

Table 1: LUMO and HOMO energies and the some corresponding Global quantum physic-chemical descriptors (GQCDs).

Quantum parameters
E_LUMO(eV)	5.971
E_HOMO(eV)	-1.135
Ionization potential (I, eV) Ionization potential (I, eV)	1.135
Electron affinity (A, eV)	-5.971
∆E gap (eV)	7.106
Dipolar moment (D)	1.583
Global hardness (η, eV)	3.553
Global softness (σ, eV^-1)	0.281
Electronegativity (χ, eV)	-2.418
Chemical potential (μ, eV)	2.418
Global electrophlicity (ɷ, eV) eV)	0.823

Figure 5: Frontiers Molecular Orbitals plots.

Chemical reactivity and stability:

Through the gap energy E_GAPthat has the value7.106eV for the Bis-AcPh, the chemical stability can be explained. So, the large value reflects the low reactivity and high stability and vice versa. In addition, the hardness, softness and chemical potential having the values 3.553eV, 0.281eV^-1 and 2.418eV respectively are other parameters that can discuss the reactivity and stability, soft molecules has a less energy of excitation compared to the hard molecule as the Bis-AcPh, which requires high energy of excitation.²⁵ The calculation of the GQCDs indicates that the μ was found 2.418eV, this value suggests according to chelation therapy, that charges transfer with the external medium may occur.²⁶ To assess the molecule’s efficiency to donate electrons to the surrounding medium, the Q_max was estimated according to the Formula:

Q_max = −μ/η

The total electronic charge Q_max proves again the donating character of the Bis-AcPh molecule.²⁷

Antioxidant activity:

Scavenging effect on DPPH radical:

Molecules designated as antioxidants are substances that generously donate electrons to offset and neutralize harmful oxidants.²⁸ DPPH assay is widely employed in determining the potential of radical scavenging activity²⁹because of its stability in the radical form along with the simplicity of the test. DPPH is a relatively stable free radical because it has the ability to either gain an electron or a proton in order to produce a stable molecule. Due to its odd electron, it exhibit an intense absorption band in the visible spectrum at 517nm. When an electron donates a proton to the DPPH radical, the absorbance of DPPH solution diminishes.²⁸ This activity has been employed to assess the capability of compound and plant extracts to act as free radical scavengers.³⁰ The antioxidant potential of Bis-AcPh against DPPH radical was investigated in respect to the 50% inhibitory concentration (IC₅₀) values. Results reveal that the molecule Bis-AcPh scavenged the DPPH radical with IC₅₀ values 459.81±0.09μg/ml meanwhile, that of the standard drug BHT antioxidant was 87.26±0.01μg/ml. These findings affirm that Bis-AcPh exhibited a moderate scavenging activity against this free radical.

Reducing power assay:

The reduction of Fe³⁺is frequently employed to measure the electron donating activity present in a given sample. In the reducing power procedure, the existence of antioxidants in the sample reduce Fe³⁺to Fe²⁺ by giving an electron, the proportion of Fe²⁺ complex in the samples can be then determined by Perl’s Prussian blue formation at 700nm. Higher absorbance value at the final reaction mixture at 700nm signifies that the compounds tested have higher reducing capacity.³¹ Our investigation showed in (Figure 7.) revealed that the IC₅₀ value of the reference compound VitC was noted as 21.91±0.48μg/ml, however the Bis-AcPh gived 459.81±0.09μg/ml.

The activities of the Bis-AcPh are much lower in term of DPPH scavenging ability and reducing power essays, this result due to it associated chemical structure that present an absence of phenolic group. Antioxidant activity is often related to the functional groups that can release or neutralise free radicals, mainly the hydroxyl (OH) often present on phenolic compounds, including polyphenols.³² The reducing properties are generally linked to the presence of reductones, which have been shown to exhibit antioxidant activity by interrupting the free radical chain through the donation of a hydrogen atom. In 1,3-bis(2-Acetylphenoxy)-2-propanol,the two aromatic rings are acetylated at the ortho position, the active hydrogen that ordinarily functions in antioxidant reactions as the phenols is replaced by the acetyl group (COCH₃). This modification slows down the possibility of the molecule to donate a hydrogen atom or electrons, which are crucial in neutralizing reactive oxygen species.

Furthermore, highly active antioxidant compounds often have double bounds and resonance structures that stabilize the radicals produced after donating electrons or hydrogen atoms. The acetylation of the two phenolic hydroxyls in Bis-AcPh disrupt this sort of resonance stability and hence, our compound has poor antioxidants properties. Generally, synthetic antioxidants are compounds containing phenolic moieties with various degree of alkyl substitution.³³ These phenolic antioxidants transform Peroxy radicals into hydroperoxides and during oxidation becoming phenoxy radicals themselves. The phenoxy radicals can intereact in different manners with another peroxy radical leading to a nonradical products.³⁴

Anti-inflammatory activity:

In vitro anti-inflammatory activity:

One of the established in vitro tests to ascertain whether a natural or synthetic compound has anti-inflammatory properties is through a protein denaturation test, as inflammation is one of the main leading of proteins denaturation.³⁵ Therefore, the ability of a compound to prevent proteins denaturation, indicate that it may have anti-inflammatory potential.³⁶ The finding of anti-inflammatory screening of the Bis-AcPh and the standard Aspirin drug by employing egg albumin denaturation test are presented in Figure 6. Our results showed that the inhibition rate increase with increasing the concentration of both the Bis-AcPh and the Aspirin. The Bis-AcPh exhibited a significant inhibition level of heat-induced protein denaturation (p<0.05) almost like the aspirin at the concentration of 2.5mg/mL with values of 95.16±4.47% for the Bis-AcPh and 99.58±0.06% for the standard aspirin drug (Figure 6). Whereas, the compound shows less activity at low concentration, the standard is almost 2 times more active than the Bis-AcPh for instance at the concentration of 0.3125mg/ml. The tests were performed in quadruplicate determinations and the results are depicted as mean±standard error of the mean (SEM).

Figure 6: Protein denaturation assay of the synthesized compound Bis-AcPh and the Aspirin drug, based on the compound concentration.

Data are stated as the mean ±SD (n=4), ns: no significant difference p < 0.05, ***: p < 0.001 in comparison with Aspirin.

In vivo anti-inflammatory activity:

Xylene-induced ear edema:

The anti-inflammatory activity was characterized by the sample’s capacity to prevent edema. Xylene-induced edema is recognized for triggering acute inflammation, marked by notable vasodilation and changes in skin edema. Mice skin exposed to the chemical xylene produces the release of inflammatory substances like histamine, serotonin, bradilkynin, and TNF-α.³⁷ In this experiment, the ability of the compound Bis-AcPh to inhibit inflammation induced by xylene was estimated using mice model. The results showed that the activity of compound is better than that of the positive drug indomethacin, which is expressed as the rate of inhibition of 81.54±6.52%. However, the inhibition rates obtained with Bis-AcPh is 86.15±2.66%. In this way, we were able to identify that the compound we obtained has certain anti-inflammatory abilities that could be applied in the treatment of inflammatory diseases.

Croton oil-induced ear edema:

The induction of ear edema by croton oil is an in vivo test frequently utilized for the examination of topical acute anti-inflammatory activity. This test is simple, effective, sensitive and quick and involves minimal number of compounds for performing assays. This model has been also extensively employed for screening and evaluating novel candidate compounds having anti-inflammatory activity, and could be used in the treatment of inflammatory infections that affect the skin.³⁸ The major irritant in croton oil 12-O-tetradecanoylphorbol-13-acetate (TPA), launches the activation of protein kinase C, this activation, in turn, induce the release of pro-inflammatory cytokines, as well as other mediators including arachidonic acid and phospholipase A2³⁹ leading to the release of platelet activation factor. This series of events triggers an increase in vascular permeability and vasodilation, migration of polymorphonuclear leukocytes, and the release of histamine and serotonine, along with a moderate synthesis of inflammatory eicosanoids, as a response to the irritation produced by the chemical.⁴⁰ Our findings revealed that topical application of croton oil on the ear of mice produced significant oedema formation. However, the bis-ketone Bis-AcPh significantly revealed low ear thickness at 6h after being subjected to croton oil, the positive control Indomethacin at a concentration of 2mg, inhibited mice ear edema by 61.02%±3.42%, while Bis-AcPh with a concentration of 2mg/ml, produced an inhibition of 71.28%±5.47%.

The results of both in vitro and in vivo anti-inflammatory experiences proved that our synthetized compound shows good anti-inflammatory activity, which could be due to its structural features that permits the compound to interact with the biological process responsible for the inflammation. The phenoxy groups in the molecule of Bis-AcPh may provide opportunity for interaction with enzymes and proteins responsible for the inflammation regulation. Rao et al.⁴¹ have indicating that the phenoxy moiety played the major roles in anti-inflammatory activity of the compounds. They have explored that the molecule [2-(4-acetylphenoxy)-9,10-dimethoxy-6,7-dihydropyrimido [6,1-a]isoquinolin-4- one; assigned as EI-03] decrease levels of a molecule known as TNF-α,that usually fuels inflammation, and boosted levels of a molecule known as IL-10,which is used to reduce inflammation. This suggests that EI-03 may be a useful drug for treating autoimmune sickness by means of reducing inflammation and restoring immune system balance. Various works have been done to analysis several bis-ketone compounds especially the α,β-insaturated carbonyls groups regarding to their anti-inflammatory potencialities. These compounds, such as curcumin, chalcones and other β-diketone have strongly biological remodeling like anti-inflammatory, and the carbonyl group is evidently very important in these activities.⁴² From the obtained results, our synthetized bis-ketone might reduce the production of inflammatory eicosanoids; a class of molecules that inhibited chemotaxis of inflammatory cells and diminished vascular permeability, contributing to the observed anti-inflammatory effect.⁴³

Cytotoxicity against red blood cells:

The hemolytic test was used to investigate the biocompatibility of the synthetic molecule Bis-AcPh. The hemolysis rate was determined by contrasting the molecule’s absorbance with that of the reference control Triton_X-100 that exhibited 100% hemolysis. The finding indicate that the hemolysis rate of the red blood cells caused by the Bis-AcPh ranged from 0.42±1.95% to 1.00±3.11% compared to the Triton_X-100. The results are shown in (Figure 7).

All drug delivery systems introcuced into the blood streams interacts with red blood cells RBCs. The in vitro erythrocyte caused hemolysis is regarded to be an easy and dependable protocol in order to estimate the compatibility of the Bis-AcPh with the blood. The hemolytic assay results in this experience indicate that even at relatively high concentration (up to 2mg/ml) our compound did not exhibit valuable hemolytic activity towards human electrolytes (RBCs). The stage of hemolysis in in-vitro investigations might be classified as“not significant” when it varies between the range 5 to 25%⁴⁴. In our investigations, we discovered that the haemolytic degree of Bis-AcPh was less than 2% in the examined concentration range. Therefore, the Bis-AcPh didn’t exhibit any noticeable hemolytic impact on human red blood cells. In addition, this is a good indicator of biocompatibility of this molecule.

Figure 7: The hemolysis rate induced on humain red blood cells by various dose of Bis-AcPh. Values are represented as the mean±SD (n = 3), ***:p< 0.001 in comparison to Triton X-100.

Somme examples of bis-ketones have been explored for their low cytotoxicity particularly in the field of pharmaceutical or biomedical applications, among them the curcumin and its derivatives. A research assessed the difference between the hemolytic activity of curcumin and nanocurcumin monstrate that evidently, nanocurcumin has a slight significantly higher affinity to RBCs compared to the curcumin, but even at this state the level of hemolysis remained was extremely low and there is no significant difference between the level of hemolysis of nanocurcumin and parent curcumin.⁴⁵

Hemolytic compound tend to be lipophilic and amphiphilic, which enhances their ability to break cell membranes. The amphiphilic character of Triton X-100 enables it to dissolve well in water based solutions as well as aqueous and hydrophobic while our compound Bis-AcPh is less soluble in water and it does not reach the amphiphilicity needs to break membranes. It might lack the right hydrophilic equilibrium necessary for breaking cell membranes similarly to Triton X-100.

CONCLUSION:

In summary, the bis-ketone Bis-AcPh has been synthesized and characterized using some spectroscopic methods. The antioxidant activity of the synthesized compound Bis-AcPh investigated by both DPPH radical-scavenging and reducing power method showed low antioxidant effect. Whereas, the compound has demonstrated anti-inflammatory activities. In addition, Assessment of toxicity demonstrated no toxic effect of Bis-AcPh against red blood cells. DFT calculations were employed to obtain the 3D molecular structure, and to investigate the electronic properties and the chemical reactivity. At the end and through the theoretical calculations a process discussing the correlation between the electronic properties and the antioxidant activity was proposed.

ACKNOWLEDGMENTS:

The authors wish to express their gratitude to the Directorate General of Scientific Research and Technological Development (DGRSDT) and the Algerian Ministry of Higher Education and Scientific Research (MESRS) for their financial support.

CONFLICT OF INTEREST:

The authors have no conflicts of interest regarding this investigation.

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Received on 08.01.2025 Revised on 05.05.2025

Accepted on 13.07.2025 Published on 08.11.2025

Available online from November 13, 2025

Research J. Pharmacy and Technology. 2025;18(11):5128-5136.

DOI: 10.52711/0974-360X.2025.00740

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