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Article

Synthesis, Antiprotozoal Activity, and Physicochemical Evaluation of Benzamido–Menadione Derivatives

by
Armin Presser
1,*,
Gregor Blaser
1,
Eva-Maria Pferschy-Wenzig
2,3,
Monica Cal
4,5,
Pascal Mäser
4,5 and
Wolfgang Schuehly
2
1
Institute of Pharmaceutical Sciences, Pharmaceutical Chemistry, University of Graz, Schubertstrasse 1, 8010 Graz, Austria
2
Institute of Pharmaceutical Sciences, Pharmacognosy, University of Graz, Beethovenstrasse 8, 8010 Graz, Austria
3
Field of Excellence BioHealth, University of Graz, 8010 Graz, Austria
4
Swiss Tropical and Public Health Institute, Kreuzstrasse 2, 4123 Allschwil, Switzerland
5
Faculty of Philosophy and Natural Sciences, University of Basel, Swiss TPH, Petersplatz 1, 4003 Basel, Switzerland
*
Author to whom correspondence should be addressed.
Int. J. Mol. Sci. 2025, 26(22), 10951; https://doi.org/10.3390/ijms262210951
Submission received: 24 October 2025 / Revised: 7 November 2025 / Accepted: 10 November 2025 / Published: 12 November 2025
(This article belongs to the Special Issue Synthetic Chemistry in Drug Discovery)
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Abstract

The naphthoquinone skeleton is known for broad biological applications and, in particular, for antiparasitic efficacy. As part of our ongoing search for new antiprotozoal naphthoquinone derivatives, we incorporated computer-aided optimization models utilizing physicochemical parameters into our approach. Herein, we report on the synthesis of 21 new benzamido–menadione and naphthoquinone derivatives via the Kochi–Anderson reaction. The antiprotozoal activity of all the synthesized compounds was evaluated against Plasmodium falciparum NF54 and Trypanosoma brucei rhodesiense STIB900. Cytotoxicity towards L6 cells was also determined, and the respective selectivity indices (SI) were calculated. Several ligand efficiency metrics, such as LLE, SILE, and FQ, were calculated, and the results were visualized in scatterplots. Almost all of the synthesized benzamido–menadione derivatives exhibited high activity against NF54 (IC50 < 1 µM), with the strongest activity and excellent selectivity observed in the 2-fluoro-5-trifluoromethylbenzamido derivative 2f (IC50 = 0.021 µM, SI = 10,000). Specific ligand efficiency metrics, such as SILE, LLE or FQ, showed a clear correlation with the corresponding antiplasmodial activities. Toxicity predictions confirmed low acute oral toxicity for most compounds, further supporting their potential as safe drug candidates. Our findings highlight the benzamido–menadione scaffold as a viable option for new antiplasmodial drugs.

1. Introduction

Malaria, the world’s most devastating parasitic disease, remains a serious health threat. In 2023, the World Health Organization (WHO) estimated 263 million cases and 597,000 deaths globally, a rise of 11 million cases from 2022, with most occurring in the African region [1,2]. Progress in the global fight against malaria has stalled slightly in recent years. Therefore, the WHO is requesting increased efforts and additional funding to support the strategic use of new tools, including vaccines and new antimalarial drugs.
Alkyl-1,4-naphthoquinones, including plumbagin, plasmodione, and phytomenadione (vitamin K1) (Figure 1), have attracted considerable attention due to their significant biological activity [3,4,5,6,7]. In particular, the redox-active compound plasmodione has emerged as a promising candidate in the search for new malaria drugs due to its multifactorial mechanism of action against different stages of the malaria parasite Plasmodium falciparum with varying degrees of drug resistance [8,9,10,11]. This compound belongs to the class of 1,4-naphthoquinones, which are recognized for their antiparasitic efficacy and broad biological applications [12,13,14,15,16]. In addition, plasmodione has demonstrated a favourable safety profile, which enables a possible human use, including for patients with G6PD deficiency [17].
In the present study, 21 plasmodione-like derivatives with a benzamido–naphthoquinone skeleton were synthesized. The strategic application of physicochemical parameters is of particular importance in this work due to their decisive role in rational drug development. Therefore, in order to assess the usability of the synthesized compounds, an analysis of various physicochemical parameters and ligand efficiency metrics was conducted, as well as toxicity studies.

2. Results and Discussion

2.1. Synthetic Chemistry

We recently reported a simple and fast preparation route for benzyl-substituted 1,4-naphthoquinone (NQ) and menadione (MD) derivatives via the Kochi–Anderson reaction [18]. Silver-catalyzed coupling between an NQ core and substituted phenylacetic acids is an efficient method for synthesizing these derivatives in a single step [9,19,20]. However, it has been demonstrated that the use of aminophenylacetic acids as coupling agents yielded only very modest results, and in addition, it requires the application of protective groups [21].
In the present study, we employed a variety of benzamido-phenylacetic acids as reactants, which were synthesized from commercially available aminophenylacetic acids and (mainly fluorinated) benzyl chlorides as protecting groups (Scheme 1), according to Billamboz et al. [22]. Based on our previous work, we first synthesized a series of para-(2a–i) and meta-substituted menadione derivatives (3a–h) in good-to-satisfactory yields (61–83%; see Scheme 1, route A).
For the preparation of the benzamido-NQ derivatives, the procedure had to be slightly modified by adjusting the amounts of the reagents used to minimize the formation of disubstituted compounds. Consequently, the para-substituted compounds 5a,b and the meta-derivatives 6a,b could be obtained in moderate yields (53–59%) (Scheme 1, route B). The synthesized compounds, together with their synthetic pathways and the resulting yields, are presented in Table 1.

2.2. Biological Evaluation

Synthesized benzamido–menadione and naphthoquinone derivatives were tested in vitro for their antiprotozoal activity against P. falciparum (NF54) in the erythrocyte stage and T. brucei rhodesiense (STIB900) in the bloodstream form. The evaluation of the compounds’ potential for toxicity was conducted using rat L6 skeletal muscle cell lines, and their corresponding selectivity index (SI) was determined based on the ratio between the cytotoxic and antiplasmodial activity of each compound (SI = IC50(L6)/IC50(parasite)).
According to the recommended hit-to-lead identification criteria [23,24,25], almost all benzamido–menadiones exhibited high activity towards NF54 (IC50 < 1 µM, Table 2) and favourable selectivity, particularly 2c, 2e, 2f, 2g, and 2i (SI = 2900–10,000). The strongest activity in the NF54 assay with excellent selectivity was exhibited by the 2-fluoro-5-trifluoromethylbenzamido derivative 2f (IC50 = 0.021 μM, SI = 10,000). In this setting, only the meta-substituted benzamido–menadiones 3a, 3d, 3g, and 3h displayed moderate antiplasmodial effects (IC50 = 1–14 µM). The benzamido–naphthoquinones (5a, 5b, 6a, and 6b) generally displayed lower antiplasmodial activity and selectivity. A certain trypanocidal effect was also observed (most notably with benzamido-NQ 5a), although it was generally much less developed.
Toxicity prediction was performed using the online platform ProTox-3.0, a web-based tool that utilizes a comprehensive database and was developed to evaluate the toxicological properties of chemical compounds using advanced machine learning algorithms [26]. These predictions indicated low acute oral toxicity for our compounds, with LD50 values of no less than 1000 mg/kg. The only negative outlier was the trifluoromethoxy derivative 2h, with a calculated oral toxicity of 57 mg/kg. Due to their presumed negligible oral toxicity [27], the development of drugs based on the benzamido–menadione scaffold could be a promising approach to retrieve new antiprotozoal agents.

2.3. Physicochemical Investigations

The hit-to-lead process (H2L) is an essential part of modern drug discovery and involves the optimization of validated hit structures into lead molecules [28,29,30]. While a lead molecule may not yet be perfect, it has structural properties as well as inherent efficacy and physicochemical characteristics that would be difficult to address in later phases of drug development. Calculated physicochemical properties, especially those falling under the rule of five (Ro5), have long played a central role in drug development, as they have been shown to be effective tools in predicting the absorption and permeation of compounds [31,32,33,34,35].
An important factor in selecting hits is to prioritize compounds with higher ligand efficiency. Of all the ligand efficiency metrics (LE, SILE, FQ, LLE, and LELP), the lipophilic ligand efficiency LLE (also known as LipE) is the most robust and widely applicable due to its correlation to enthalpy [36,37,38]. A selection of important ligand efficiency metrics and multi-parameter scores, along with their calculation method, is shown in Table 3.
The size-independent ligand efficiency (SILE) [39] and fit quality (FQ) [40] scores have been developed to identify drug candidates across a wide range of ligand sizes, thereby facilitating the discovery of ligands with good ligand efficiency or even ligands with exceptional efficiency.
However, it has been shown that setting ambitious scores for these metrics is unwise, as they are target-dependent [29]. The recommended strategy is to focus on maximizing the pivotal LLE and prioritizing series with higher average values. Due to their significance in the drug discovery process, we calculated the most important physicochemical parameters, ligand efficiency indices, and multi-parameter scores of our tested compounds and listed them in Tables S1 and S2 in the Supplementary Material.

2.4. Structure-Activity Relationships (SAR) of the Antiplasmodial Activity

The vast majority of the compounds examined in this study complied with the established recommendations of Ro5 [41], AB-MPS [42], and PFI [43]. However, some argue that focusing too much on these guidelines hinders the development of new drug leads [44]. Therefore, this study focuses on ligand efficiency metrics as the most important tool for lead optimization and controlling lipophilicity in relation to efficacy.
As in our previous work [18], we found that some of the calculated ligand efficiency metrics, especially size-normalized SILE and FQ, correlated highly with the observed antiplasmodial activity (e.g., SILEP.f., ρ = −0.99; FQP.f., ρ = −0.99). Spearman’s rho (ρ) used for this purpose is robust against outliers and does not require normally distributed data, as extreme values are converted into ranks, thereby reducing their disproportionate influence on the calculation. The observed Spearman correlation for SILE and FQ confirms that hit compounds can be more accurately identified by adjusting their lipophilicity relative to their molecular size. It is noteworthy to mention that alternative size-independent ligand efficiency metrics, such as the ligand efficiency lipophilicity price (LELP) [45] and the Astex lipophilicity ligand efficiency (LLEAT) [46], exhibited no substantial correlation with the efficacy of our compounds.
Examination of the ligand efficiency metrics of our compounds in an LLE/SILE scatterplot revealed clear correlations between compounds with similar structural elements and corresponding antiplasmodial activities (Figure 2 and Figure S1 in the Supplementary Materials). Here, the activity of para-substituted benzamido–menadiones is particularly evident, as all these compounds are situated in the recommended upper-right quadrant (LLE > 1.1, SILE > 2.2) and demonstrate remarkable antiplasmodial activity (pIC50 ≥ 6.9). Among the para-MD derivatives, the multifluorinated derivative 2i (pIC50 of 7.6) is particularly outstanding; only compounds 2a and 2d exhibited slightly lower values (with a pIC50 of 6.2 and 6.5, respectively). In contrast, the NQ derivatives synthesized in our study (5a, 5b, 6a, and 6b) generally showed reduced activity (pIC50 ≤ 5.7) and were predominantly located in the lower-left region of the LLE/SILE scatterplot.
The trajectories of selected compounds in the LLE and SILE space are also shown in Figure 2, providing an excellent opportunity to evaluate the optimization process in drug development. For instance, an alteration from meta- to para-substituted derivatives (3d → 2d, 3f → 2f, 3g → 2g, 3h → 2h) has been observed to result in a substantial enhancement in effectiveness, as indicated by an explicit migration toward the north-eastern quadrant of the LLE-SILE plot. In contrast, the removal of the methyl group from the menadione backbone results in a substantial reduction in its antiplasmodial effect (2a → 5a, 2b → 5b, 3b → 6b).
Incidentally, this behaviour is not only limited to the LLE/SILE plane; the LLE/FQ alternative performed is almost identical to the LLE/SILE plot (see Figure S2 in the Supplementary Material), suggesting its potential application in a comparable manner. These results underscore the potential of the LLE/SILE, as well as the LLE/FQ plot, to serve as valuable tools regarding the effectiveness of different functional groups and moieties in the context of drug development.
In light of the prevailing consensus that LLE is the most robust and widely used metric for assessing quality in drug discovery [36], we also correlated this key parameter with the selectivity of the tested compounds as log SI (Figure 3).
The remarkable capability of p-substituted MDs is further evidenced by the LLE/log SI scatterplot, which exhibited a clear clustering of the evaluated compounds according to lipophilic efficiency and selectivity. With the exception of 2a (log SI = 2.7) and 2d (log SI = 2.6), the log SI values are > 3.2 in all cases, indicating that these compounds can be found in the desired upper-right section of the LLE/log SI chart. The meta-MD derivatives are situated in the central region of the scatterplot (with the exception of 3g), while 3g and all of the NQs exhibit less favourable prospects and are located in the southwestern part of the plane.
Abad-Zapatero et al. proposed an alternative approach to guide the transition from hit to lead, which involved the implementation of the binding efficacy index (BEI), an extension of the ligand efficiency (LE), and the surface-binding efficacy index (SEI) [47]. This concept was enhanced by the introduction of the Atlas-CBS system, which enables efficient lead optimization and prioritization by mapping the distribution of various compounds in an nBEI-NSEI plane [48,49,50]. This pair of variables is ideal for identifying the ‘direction’ of the optimization path provided by the corresponding scaffold, which corresponds to the slope given by NPOL (N + O count). It also allows for identifying likely drug candidates in the most probable ‘efficiency region’ situated in the north-eastern part of the plane. Furthermore, this approach facilitates the establishment of a direct graphical visualization of the guidelines proposed by Lipinski and colleagues, as outlined in Ro5 [50]. This provides further insight into the Ro5, in which the potency of the compounds is not considered.
The application of the Atlas CBS system to our compounds corresponds remarkably well with the LLE/SILE plot described above. In this instance as well, the para-MD derivatives (again with the exception of 2a and 2d) have been identified as the most efficient compounds in terms of size and polarity and are located in the upper-right corner of the plane (Figure 4 and Figure S3 in the Supplementary Materials). Consequently, these compounds are regarded as promising candidates for further drug development.
The concept of trajectory mapping can also be used very effectively in the NSEI/nBEI scatterplot to monitor the optimization process in drug discovery. Again, the switch from meta- to para-substituted derivatives (3e → 2e, 3f → 2f, 3g → 2g, 3h → 2h) resulted in a substantial surge in activity, as demonstrated by a discernible shift toward the preferred north-east direction depicted in the NSEI/nBEI correlation plot. In contrast, the activity of all NQ derivatives was less promising, which is also clearly reflected in the NSEI/nBEI chart.

3. Materials and Methods

3.1. Chemistry

3.1.1. General Information

All reagents and solvents were obtained from Merck and Fluorochem Ltd. Moisture-sensitive reactions were performed under an inert argon atmosphere. Each reaction was monitored by TLC on Merck TLC plates (silica gel 60 F254 0.2 mm, 200 × 200 mm) and detected at 254 nm. All reaction products were purified by flash column chromatography on silica gel 60 (Merck, Darmstadt, Germany, 70–230 mesh, pore diameter 60 Å) unless otherwise stated. Purity and homogeneity of the final compounds were checked by TLC and high-resolution mass spectrometry. Melting points were determined using a digital melting point apparatus (Electrothermal IA 9200, Thermo Fisher Scientific, Birmingham, UK).
The structures of all synthesized derivatives were determined by 1D and 2D NMR spectroscopy on a Bruker Avance Neo 400 MHz instrument (at 298 K) using 5 mm tubes. Chemical shifts were expressed in δ (ppm) using either tetramethylsilane (TMS) or the 13C signal of the solvents (CDCl3 δ 77.04 ppm, DMSO-d6 δ 39.45 ppm) as the internal standard. Accurate structural elucidation was confirmed by 1H, 1H- and 1H, 13C-correlation spectra (COSY, HSQC, HMBC, H2BC). 1H and 13C resonances were numbered according to the formulae (see Supplementary Material), and signals marked with an asterisk are interchangeable.
ESI and APCI mass spectra were acquired by analyzing sample solutions on an Ultimate 3000 HPLC with a Q Exactive™ Hybrid Quadrupole-Orbitrap™ mass spectrometer equipped with a heated ESI II source or an APCI source (Thermo Fisher Scientific) in positive or negative ionization mode.

3.1.2. General Synthetic Procedure for the Preparation of Benzamido-Phenylacetic Acids

The aminophenylacetic acid (1 mmol) was dissolved in 2.0 M KOH (10 mL). Then, the benzoyl chloride derivative (1.1 equiv.) was added dropwise, and the solution was stirred at room temperature until TLC showed complete consumption of the starting material (18–24 h). After acidifying to pH 4.0 with 2.0 M aqueous HCl, the mixture was extracted three times with EtOAc (50 mL). The combined organic layers were washed with H2O, dried over Na2SO4, and concentrated in vacuo to yield the crude products as off-white solids, which were used without further purification.

3.1.3. General Synthetic Procedure for Benzamido–Menadiones 2a–i and 3a–h (Route A)

Menadione (1.0 mmol) was added to a stirred solution of the respective benzamido-phenylacetic acid derivative (1.4 equiv.) in CH3CN (9 mL) and H2O (3 mL) and heated to 85 °C. AgNO3 (0.35 equiv.) was added first and then (NH4)2S2O8 (1.3 equiv.) dissolved in 4 mL CH3CN/H2O (3:1) dropwise over a period of 5 min. Stirring at 85 °C was continued until TLC showed complete consumption of the starting material (3–6 h). After cooling to ambient temperature, the mixture was extracted three times with CH2Cl2 (30 mL). The combined organic layers were washed with H2O, dried over Na2SO4, and concentrated in vacuo to give a residue, which was purified by flash chromatography as detailed below.
N-{4-[(3-Methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}benzamide (2a). Compound 2a was obtained after stirring for 3 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 79%; Rf = 0.53 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 195–196 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.18 (s, 1H, NH), 8.01 (m, 2H, H-5, H-8), 7.92 (m, 2H, H-2″,H-6″), 7.85 (dd, J = 6.0, 3.4 Hz, 2H, H-6, H-7), 7.66 (d, J = 8.3 Hz, 2H, H-3′, H-5′), 7.57 (m, 1H, H-4″), 7.52 (d, J = 7.5 Hz, 2H, H-3″, H-5″), 7.21 (d, J = 8.3 Hz, 2H, H-2′, H-6′), 3.96 (s, 2H, H-9), 2.17 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.7 (C-4), 184.1 (C-1), 165.3 (CO(NH)), 144.5 * (C-2), 144.1 * (C-3), 137.3 (C-4′), 134.9 (C-1″), 133.9 (C-7), 133.8 (C-6), 133.3 (C-1′), 131.7 (C-4a), 131.4 (C-8a, C-4″), 128.4 (C-2′, C-6′), 128.3 (C-3″, C-5″), 127.6 (C-2″, C-6″), 125.8 (C-5, C-8), 120.5 (C-3′, C-5′), 31.1 (C-9), 12.9 (CH3) ppm; HRMS (ESI) calcd. for C25H20NO3 [M + H]+ = 382.1438; found: 382.1433.
N-{4-[(3-Methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-4-(trifluoromethyl)benzamide (2b). Compound 2b was obtained after stirring for 4 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 72%; Rf = 0.47 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 210–211 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.40 (s, 1H, NH), 8.11 (d, J = 8.1 Hz, 2H, H-2″, H-6″), 8.01 (m, 2H, H-5, H-8), 7.89 (d, J = 8.1 Hz, 2H, H-3″, H-5″), 7.84 (m, 2H, H-6, H-7), 7.66 (d, J = 8.4 Hz, 2H, H-3′, H-5′), 7.22 (d, J = 8.4 Hz, 2H, H-2′, H-6′), 3.97 (s, 2H, H-9), 2.17 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.7 (C-4), 184.1 (C-1), 164.1 (CO(NH)), 144.4 * (C-2), 144.2 * (C-3), 138.7 (C-1″), 136.9 (C-4′), 133.9 * (C-6), 133.8 * (C-7), 133.7 (C-1′), 131.6 * (C-8a), 131.4 * (C-4a), 131.1 (q, 2JC,F = 32.0 Hz, C-4″), 128.5 (C-2′, C-6′, C-2″, C-6″), 125.9 * (C-5), 125.8 * (C-8), 125.3 (q, 3JC,F = 3.7 Hz, C-3″, C-5″), 123.8 (q, 1JC,F = 272.5 Hz, CF3), 120.6 (C3′, C-5′), 31.1 (C-9), 12.9 (CH3) ppm; HRMS (ESI) calcd. for C26H17F3NO3 [M − H]– = 448.1161; found: 448.1173.
2-Fluoro-N-{4-[(3-methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-4-(trifluoromethyl)benzamide (2c). Compound 2c was obtained after stirring for 4.5 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 81%; Rf = 0.43 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 181–182 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.54 (s, 1H, NH), 8.01 (m, 2H, H-5, H-8), 7.85 (m, 2H, H-3″, H-6″), 7.84 (m, 2H, H-6, H-7), 7.71 (d, J = 8.1 Hz, 1H, H-5″), 7.60 (d, J = 8.3 Hz, 2H, H-3′, H-5′), 7.23 (d, J = 8.3 Hz, 2H, H-2′, H-6′), 3.97 (s, 2H, H-9), 2.16 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.6 (C-4), 184.1 (C-1), 161.3 (CO(NH)), 158.5 (d, 1JC,F = 251.1 Hz, C-2″), 144.4 * (C-2), 144.2 * (C-3), 136.6 (C-4′), 133.9 (C-1′), 133.9 * (C-6), 133.8 * (C-7), 132.1 (qd, 2JC,F = 33.0 Hz, 3JC,F = 8.3 Hz, C-4″), 131.6 * (C-4a), 131.4 * (C-8a), 131.0 (d, 3JC,F = 3.7 Hz, C-6″), 128.9 (d, 2JC,F = 16.6 Hz, C-1″), 128.7 (C-2′, C-6′), 125.9 * (C-5), 125.8 * (C-8), 123.0 (dd, 1JC,F = 272.8 Hz, 4JC,F = 2.5 Hz, CF3), 121.5 (quint, 4JC,F = 3.7 Hz, C-5″), 119.9 (C-3′, C-5′), 113.8 (dq, 2JC,F = 25.6 Hz, 3JC,F = 3.8 Hz, C-3″), 31.1 (C-9), 12.9 (CH3) ppm; HRMS (ESI) calcd. for C26H16F4NO3 [M − H]– = 466.1066; found: 466.1073.
4-Fluoro-N-{4-[(3-methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}benzamide (2d). Compound 2d was obtained after stirring for 4 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 76%; Rf = 0.49 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 198–200 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.20 (s, 1H, NH), 8.01 (m, 4H, H-5, H-8, H-2″, H-6″), 7.85 (m, 2H, H-6, H-7), 7.65 (d, J = 8.5 Hz, 2H, H-3′, H-5′), 7.34 (m, 2H, H-3″, H-5″), 7.21 (d, J = 8.5 Hz, 2H, H-2′, H-6′), 3.97 (s, 2H, H-9), 2.17 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.7 (C-4), 184.1 (C-1), 164.8 (d, 1JC,F = 250.4 Hz, C-4″), 164.2 (CO(NH)), 144.5 * (C-2), 144.2 * (C-3), 137.2 (C-4′), 133.8 (C-6, C-7), 133.4 (C-1′), 132.0 (d, 3JC,F = 9.7 Hz, C-2″, C-6″), 131.7 * (C-4a), 131.4 * (C-8a), 128.4 (C-2′, C-6′), 127.3 (d, 4JC,F = 2.9 Hz, C-1″), 125.9 * (C-5), 125.8 * (C-8), 120.6 (C-3′, C-5′), 115.5 (d, 2JC,F = 21.9 Hz, C-3″, C-5″), 31.1 (C-9), 12.9 (CH3) ppm; HRMS (ESI) calcd. for C25H19FNO3 [M + H]+ = 400.1349; found: 400.1339.
4-Fluoro-N-{4-[(3-methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-3-(trifluoromethyl)benzamide (2e). Compound 2e was obtained after stirring for 4.5 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 65%; Rf = 0.40 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 170–171 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.40 (s, 1H, NH), 8.31 (m, 2H, H-2″, H-6″), 8.00 (m, 2H, H-5, H-8), 7.83 (m, 2H, H-6, H-7), 7.68 (t, J = 9.9 Hz, 1H, H-5″), 7.64 (d, J = 8.5 Hz, 2H, H-3′, H-5′), 7.23 (d, J = 8.5 Hz, 2H, H-2′, H-6′), 3.96 (s, 2H, H-9), 2.16 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.6 (C-4), 184.1 (C-1), 162.8 (CO(NH)), 160.5 (dq, 1JC,F = 258.5, 3JC,F = 2.2 Hz, C-4″), 144.4 * (C-2), 144.2 * (C-3), 136.8 (C-4′), 134.9 (d, 3JC,F = 9.9 Hz, C-6″), 133.8 (C-6, C-7, C-1′), 131.6 (d, 4JC,F = 3.5 Hz, C-1″), 131.6 * (C-4a), 131.4 * (C-8a), 128.5 (C-2′, C-6′), 126.8 (qd, 3JC,F = 4.5, 2.0 Hz, C-2″), 125.9 * (C-5), 125.8 * (C-8), 122.3 (qd, 1JC,F = 272.5, 3JC,F = 1.1 Hz, CF3), 120.7 (C-3′, C-5′), 117.4 (d, 2JC,F = 21.0 Hz, C-5″), 116.5 (qd, 2JC,F = 32.7, 12.7 Hz, C-3″), 31.1 (C-9), 12.9 (CH3) ppm; HRMS (ESI) calcd. for C26H16F4NO3 [M − H]– = 466.1066; found: 466.1073.
2-Fluoro-N-{4-[(3-methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-5-(trifluoromethyl)benzamide (2f). Compound 2f was obtained after stirring for 5 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 62%; Rf = 0.42 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 179–180 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.53 (s, 1H, NH), 8.01 (m, 3H, H-5, H-8, H-6″), 7.96 (m, 1H, H-4″), 7.84 (m, 2H, H-6, H-7), 7.60 (d, J = 8.6 Hz, 2H, H-3′, H-5′), 7.59 (t, J = 9.1 Hz, 1H, H-3″), 7.23 (d, J = 8.6 Hz, 2H, H-2′, H-6′), 3.97 (s, 2H, H-9), 2.16 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.6 (C-4), 184.1 (C-1), 161.0 (CO(NH)), 160.8 (dq, 1JC,F = 255.9, 5JC,F = 1.2 Hz, C-2″), 144.4 * (C-2), 144.2 * (C-3), 136.6 (C-4′), 133.9 (C-1′), 133.9 * (C-6), 133.8 * (C-7), 131.6 * (C-4a), 133.4 * (C-8a), 129.6 (C-4″), 128.7 (C-2′, C-6′), 127.3 (p, 3JC,F = 3.9 Hz, C-6″), 125.9 (d, 2JC,F = 17.0 Hz, C-1″), 125.9 * (C-5), 125.8 * (C-8), 125.3 (qd, 2JC,F = 32.8, 4JC,F = 3.4 Hz, C-5″), 123.5 (q, 1JC,F = 272.1 Hz, CF3), 120.0 (C-3′, C-5′), 117.6 (d, 2JC,F = 23.6 Hz, C-3″), 31.1 (C-9), 12.9 (CH3) ppm; HRMS (APCI) calcd. for C26H18F4NO3 [M + H]+ = 468.1217; found: 468.1201.
4-Fluoro-N-{4-[(3-methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-2-(trifluoromethyl)benzamide (2g). Compound 2g was obtained after stirring for 4 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 81%; Rf = 0.43 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 181–182 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.50 (s, 1H, NH), 8.01 (m, 2H, H-5, H-8), 7.84 (m, 2H, H-6, H-7), 7.76 (m, 2H, H-3″, H-6″), 7.66 (td, J = 8.5, 2.6 Hz, 1H, H-5″), 7.56 (d, J = 8.3 Hz, 2H, H-3′, H-5′), 7.21 (d, J = 8.3 Hz, 2H, H-2′, H-6′), 3.96 (s, 2H, H-9), 2.17 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.7 (C-4), 184.1 (C-1), 164.4 (CO(NH), 161.8 (d, 1JC,F = 248.5 Hz, C-4″), 144.4 * (C-2); 144,2 * (C-3), 136.9 (C-4′), 133.9 * (C-6), 133.8 * (C-7), 133.7 (C-1′), 132.8 (C-1″), 131.6 * (C-4a), 131.4 * (C-8a), 131.3 (d, 3JC,F = 8.6 Hz, C-6″), 128.6 (C-2′, C-6′), 128.1 (qd, 2JC,F = 32.6, 3JC,F = 8.1 Hz, C-2″), 125.9 * (C-5), 125.8 * (C-8), 122.8 (qd, 1JC,F = 274.0, 4JC,F = 2.6 Hz, CF3), 119.8 (C-3′, C-5′), 119.4 (d, 2JC,F = 21.2 Hz, C-5″), 114.0 (dq, 2JC,F = 25.5, 3JC,F = 4.8 Hz, C-3″), 31.1 (C-9), 12.9 (CH3) ppm; HRMS (ESI) calcd. for C26H16F4NO3 [M − H]− = 466.1066; found: 466.1073.
N-{4-[(3-Methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-3-(trifluoromethoxy)benzamide (2h). Compound 2h was obtained after stirring for 4 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 75%; Rf = 0.50 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 115–116 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.32 (s, 1H, NH), 8.01 (m, 2H, H-5, H-8), 7.97 (m, 1H, H-6″), 7.87 (s, 1H, H-2″), 7.83 (m, 2H, H-6, H-7), 7.66 (m, 3H, H-3′, H-5′, H-5″), 7.58 (m, 1H, H-4″), 7.22 (d, J = 8.3 Hz, 2H, H-2′, H-6′), 3.96 (s, 2H, H-9), 2.16 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.6 (C-4), 184.1 (C-1), 163.6 (CO(NH)), 148.3 (q, 3JC,F = 1.9 Hz, C-3″), 144.4 * (C-2), 144.2 * (C-3), 136.9 (C-4′), 133.8 (C-6, C-7), 133.7 (C-1`), 133.0 (C-1″), 130.9 (C-5″), 131.6 * (C-4a), 131.4 * (C-8a), 128.5 (C-2′, C-6′), 126.7 (C-6″), 125.9 * (C-5), 125.8 * (C-8), 123.9 (C-4″), 120.7 (C-3′, C-5′), 120.1 (C-2″), 120.0 (q, 1JC,F = 256.8 Hz, OCF3), 31.1 (C-9), 12.9 (CH3) ppm HRMS (ESI) calcd. for C26H17F3NO4 [M − H]− = 464.1110; found: 464.1117.
2,4,5-Trifluoro-N-{4-[(3-methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}benzamide (2i). Compound 2i was obtained after stirring for 5 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 61%; Rf = 0.45 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 175–177 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.41 (s, 1H, NH), 8.00 (m, 2H, H-5, H-8), 7.84 (m, 2H, H-6, H-7), 7.81 (m, 1H, H-6″), 7.72 (td, J = 10.3, 6.4 Hz, 1H, H-3″), 7.58 (d, J = 8.6 Hz, 2H, H-3′, H-5′), 7.21 (d, J = 8.6 Hz, 2H, H-2′, H-6′), 3.96 (s, 2H, H-9), 2.15 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.6 (C-4), 184.1 (C-1), 160.4 (CO(NH)), 154.6 (ddd, 1JC,F = 248.4, 3JC,F = 10.8, 4JC,F = 2.4 Hz, C-2″), 150.4 (ddd, 1JC,F = 252.4, 2JC,F = 14.1, 3JC,F = 13.0 Hz, C-4″), 145.8 (ddd, 1JC,F = 243.9, 2JC,F = 12.7, 4JC,F = 3.6 Hz, C-5″), 144.4 * (C-2), 144.2 * (C-3), 136.7 (C-4′), 133.9 * (C-6), 133.8 *(C-7), 133.8 (C-1′), 131.6 * (C-4a), 131.4 * (C-8a), 128.6 (C-2′, C-6′), 125.9 * (C-5), 125.8 * (C-8), 121.6 (ddd, 2JC,F = 17.5, 3JC,F = 5.0, 4JC,F = 4.2 Hz, C-1″), 120.0 (C-3′, C-5′), 118.0 (ddd, 2JC,F = 20.6, 3JC,F = 4.5, 1.5 Hz, C-6″), 106.8 (dd, 2JC,F = 29.1, 21.5 Hz, C-3″), 31.1 (C-9), 12.9 (CH3) ppm; HRMS (APCI) calcd. for C25H17F3NO3 [M + H]+ = 436.1155; found: 436.1136.
N-{3-[(3-Methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}benzamide (3a). Compound 3a was obtained after stirring for 4 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 72%; Rf = 0.53 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 190–191 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.15 (s, 1H, NH), 8.03 (m, 2H, H-5, H-8), 7.90 (m, 2H, H-2″, H-6″), 7.86 (m, 2H, H-6, H-7), 7.69 (dt, J = 8.0, 1.6 Hz, 1H, H-4′), 7.56 (m, 1H, H-4″), 7.55 (m, 1H, H-2′), 7.50 (m, 2H, H-3″, H-5″), 7.26 (t, J = 7.7 Hz, 1H, H-5′), 7.01 (ddd, J = 7.7, 1.9, 1.0 Hz, 1H, H-6′), 4.00 (s, 2H, H-9), 2.19 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.7 (C-4), 184.0 (C-1), 165.5 (CO(NH)), 144.4 * (C-2), 144.3 * (C-3), 139.3 (C-3′), 138.6 (C-1′), 134.9 (C-1″), 134.0 (C-6, C-7), 131.6 * (C-4a), 131.4 *(C-8a), 131.4 (C-4″), 128.7 (C-5′), 128.3 (C-3″, C-5″), 127.6 (C-2″, C-6″), 126.0 * (C-5), 125,9 * (C-8), 123.8 (C-6′), 119.8 (C-2′), 118.1 (C-4′), 31.7 (C-9), 13.0 (CH3) ppm; HRMS (ESI) calcd. for C25H20NO3 [M + H]+ = 382.1443; found: 382.1432.
N-{3-[(3-Methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-4-(trifluoromethyl)benzamide (3b). Compound 3b was obtained after stirring for 6 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 72%; Rf = 0.47 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 207–208 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.36 (s, 1H, NH), 8.09 (d, J = 8.1 Hz, 2H, H-2″, H-6″), 8.02 (m, 2H, H-5, H-8), 7.87 (d, J = 8.5 Hz, 2H, H-3″, H-5″), 7.85 (m, 2H, H-6, H-7), 7.70 (d, J = 7.9 Hz, 1H, H-4′), 7.54 (s, 1H, H-2′), 7.28 (t, J = 7.9 Hz, 1H, H-5′), 7.04 (d, J = 7.9 Hz, 1H, H-6′), 4.01 (s, 2H, H-9), 2.19 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.6 (C-4), 183.9 (C-1), 164.3 (CO(NH)), 144.4 (C-2, C-3), 139.0 (C-3′), 138.7 (q, 5JC,F = 1.3 Hz, C-1″), 133.9 (C-6, C-7), 131.6 * (C-4a), 131.4 * (C-8a), 131.2 (q, 2JC,F = 32.0 Hz, C-4″), 128.7 (C-5′), 128.5 (C-2″, C-6″), 126.0 * (C-5), 125.9 * (C-8), 125.3 (q, 3JC,F = 3.9 Hz, C-3″, C-5″), 124.2 (C-6′), 123.8 (q, 1JC,F = 272.4 Hz, CF3), 119.8 (C-2′), 118.2 (C-4′), 31.7 (C-9), 13.0 (CH3) ppm; HRMS (ESI) calcd. for C26H17F3NO3 [M − H]− = 448.1161; found: 448.1168.
2-Fluoro-N-{3-[(3-methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-4-(trifluoromethyl)benzamide (3c). Compound 3c was obtained after stirring for 6 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 69%; Rf = 0.49 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 177–178 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.51 (s, 1H, NH), 8.02 (m, 2H, H-5, H-8), 7.85 (m, 4H, H-6, H-7, H-3″, H-6″), 7.70 (d, J = 8.0 Hz, 1H, H-5″), 7.63 (d, J = 7.9 Hz, 1H, H-4′), 7.46 (s br, 1H, H-2′), 7.28 (t, J = 7.9 Hz, 1H, H-5′), 7.04 (d, J = 7.9 Hz, 1H, H-6′), 4.00 (s, 2H, H-9), 2.18 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.6 (C-4), 183.9 (C-1), 161.4 (CO(NH)), 158.5 (d, 1JC,F = 251.1 Hz, C-2″), 144.4 * (C-2), 144.3 * (C-3), 138.9 (C-1′), 138.7 (C-3′), 134.0 * (C-6), 133.9 * (C-7), 132.1 (qd, 2JC,F = 33.0, 3JC,F = 8.1 Hz, C-4″), 131.6 * (C-4a), 131.4 * (C-8a), 131.0 (d, 3JC,F = 3.4 Hz, C-6″), 128.9 (C-5′), 128.9 (d, 2JC,F = 15.5 Hz, C-1″), 126.0 * (C-5), 125.9 * (C-8), 124.3 (C-6′), 123.0 (qd, 1JC,F = 272.8 Hz, CF3), 121.4 (C-5″), 119.1 (C-2′), 117.5 (C-4′), 113.7 (dq, 2JC,F = 25.7 Hz, 3JC,F = 3.9 Hz, C-3″), 31.6 (C-9), 13.0 (CH3) ppm; HRMS (ESI) calcd. for C26H16F4NO3 [M − H]− = 466.1066; found: 466.1072.
4-Fluoro-N-{3-[(3-methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}benzamide (3d). Compound 3d was obtained after stirring for 4 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 71%; Rf = 0.49 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 199–200 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.16 (s, 1H, NH), 8.02 (m, 2H, H-5, H-8), 7.97 (m, 2H, H-2″, H-6″), 7.85 (m, 2H, H-6, H-7), 7.67 (dt, J = 7.9, 1.9 Hz, 1H, H-4′), 7.53 (t, J = 1.9 Hz, 1H, H-2′), 7.33 (m, 2H, H-3″, H-5″), 7.25 (t, J = 7.9 Hz, 1H, H-5′), 7.00 (dt, J = 7.9, 1.9 Hz, 1H, H-6′), 3.99 (s, 2H, H-9), 2.18 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.6 (C-4), 183.9 (C-1), 164.4 (CO(NH)), 163.9 (d, 1JC,F = 249.1 Hz, C-4″), 144.4 * (C-2), 144.3 * (C-3), 139.2 (C-3′), 138.6 (C-1′), 133.9 (C-6, C-7), 131.6 * (C-4a), 131.4 * (C-8a), 131.3 (d, 4JC,F = 2.9 Hz, C-1″), 130.3 (d, 3JC,F = 9.0 Hz, C-2″, C-6″), 128.7 (C-5′), 126.0 * (C-5), 125.9 * (C-8), 123.9 (C-6′), 119.8 (C-2′), 118.2 (C-4′), 115.2 (d, 2JC,F = 21.8 Hz, C-3″, C-5″), 31.7 (C-9), 13.0 (CH3) ppm; HRMS (ESI) calcd. for C25H17FNO3 [M − H]− = 398.1192; found: 398.1199.
4-Fluoro-N-{3-[(3-methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-3-(trifluoromethyl)benzamide (3e). Compound 3e was obtained after stirring for 4.5 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 83%; Rf = 0.37 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 168–169 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.38 (s, 1H, NH), 8.29 (m, 2H, H-2″, H-6″), 8.02 (m, 2H, H-5, H-8), 7.85 (m, 2H, H-6, H-7), 7.65 (m, 2H, H-4′, H-5″), 7.53 (t, J = 1.9 Hz, 1H, H-2′), 7.28 (t, J = 7.9 Hz, 1H, H-5′), 7.03 (m, 1H, H-6′), 4.01 (s, 2H, H-9), 2.19 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.6 (C-4), 184.0 (C-1), 163.0 (CO(NH)), 160.5 (dq, 1JC,F = 258.2, 3JC,F = 2.4 Hz, C-4″), 144.4 (C-2, C-3), 138.9 (C-3′), 138.7 (C-1′), 135.0 (d, 3JC,F = 9.7 Hz, C-6″), 134.0 * (C-6), 133.9 * (C-7), 131.7 (d, 4JC,F = 3.5 Hz, C-1″), 131.6 * (C-4a), 131.4 * (C-8a), 128.7 (C-5′), 126.9 (qd, 3JC,F = 4.6, 2.0 Hz, C-2″), 126.0 * (C-5), 125.9 * (C-8), 124.2 (C-6′), 122.3 (dq, 1JC,F = 272.5, 3JC,F = 1.1 Hz, CF3), 120.0 (C-2′), 118.4 (C-4′), 117.4 (d, 2JC,F = 20.8 Hz, C-5″), 116,4 (qd, 2JC,F = 32.7, 12.7 Hz, C-3″), 31.7 (C-9), 13.0 (CH3) ppm; HRMS (ESI) calcd. for C26H16F4NO3 [M − H]− = 466.1066; found: 466.1075.
2-Fluoro-N-{3-[(3-methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-5-(trifluoromethyl)benzamide (3f). Compound 3f was obtained after stirring for 5 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 62%; Rf = 0.48 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 173–175 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.50 (s, 1H, NH), 8.02 (m, 3H, H-5, H-6″, H-8), 7.96 (m, 1H, H-4″), 7.84 (m, 2H, H-6, H-7), 7.62 (m, 1H, H-4′), 7.58 (t, J = 9.1 Hz, 1H, H-3″), 7.47 (t, J = 1.8 Hz, 1H, H-2′), 7.28 (t, J = 7.9 Hz, 1H, H-5′), 7.04 (dt, J = 7.9, 1.8 Hz, 1H, H-6′), 4.00 (s, 2H, H-9), 2.18 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.6 (C-4), 183.9 (C-1), 161.2 (CO(NH)), 160.8 (d, 1JC,F = 255.1 Hz, C-2″), 144.3 (C-2, C-3), 138.8 (C-1′), 138.7 (C-3′), 134.0 (C-6, C-7), 131.6 * (C-4a), 131.4 * (C-8a), 129.6 (m, C-4″), 128.9 (C-5′), 127.3 (p, 3JC,F = 3.9 Hz, C-6″), 126.0 * (C-5), 125. 9 * (C-8), 125.9 (d, 2JC,F = 17.3 Hz, C-1″), 125.3 (qd, 2JC,F = 32.6, 4JC,F = 3.3 Hz, C-5″), 124.3 (C-6′), 123.5 (q, 1JC,F = 272.2 Hz, CF3), 119.2 (C-2′), 117.6 (C-4′), 117.5 (d, 2JC,F = 23.3 Hz, C-3″), 31.7 (C-9), 13.0 (CH3) ppm; HRMS (APCI) calcd. for C26H18F4NO3 [M + H]+ = 468.1217; found: 468.1202.
4-Fluoro-N-{3-[(3-methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-2-(trifluoromethyl)benzamide (3g). Compound 3g was obtained after stirring for 6 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 81%; Rf = 0.43 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 179–180 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.46 (s, 1H, NH), 8.01 (m, 2H, H-5, H-8), 7.84 (m, 2H, H-6, H-7), 7.76 (m, 1H, H-3″), 7.75 (m, 1H, H-6″), 7.63 (m, 1H, H-5″), 7.62 (dd, J = 8.3, 2.3 Hz, 1H, H-4″), 7.41 (t, J = 1.9 Hz, 1H, H-2′), 7.26 (t, J = 7.9 Hz, 1H, H-5′), 7.01 (d, J = 7.9 Hz, 1H, H-6′), 3.99 (s, 2H, H-9), 2.17 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.6 (C-4), 183.9 (C-1), 164.5 (CO(NH)), 161.8 (d, 1JC,F = 248.6 Hz, C-4″), 144.4 * (C-2), 144.3 * (C-3), 139.0 (C-3′), 138.8 (C-1′), 134.0 * (C-6), 133.9 * (C-7), 132.7 (m, C-1″), 131.6 * (C-4a), 131.4 * (C-8a), 131.3 (d, 3JC,F = 8.7 Hz, C-6″), 128.9 (C-5′), 128.1 (qd, 2JC,F = 32.6, 3JC,F = 8.1 Hz, C-2″), 126.0 * (C-5), 125.9 * (C-8), 124.0 (C-6′), 122.7 (qd, 1JC,F = 274.1, 4JC,F = 2.6 Hz, CF3), 119.4 (d, 2JC,F = 21.2 Hz, C-5″), 119.1 (C-2′), 117.4 (C-4′), 114.0 (dq, 2JC,F = 25.5, 3JC,F = 5.0 Hz, C-3″), 31.6 (C-9), 13.0 (CH3) ppm; HRMS (ESI) calcd. for C26H16F4NO3 [M − H]− = 466.1066; found: 466.1071.
N-{3-[(3-Methyl-1,4-dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-3-(trifluoromethoxy)benzamide (3h). Compound 3h was obtained after stirring for 4 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Yellow solid; yield: 78%; Rf = 0.41 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 116–117 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.29 (s, 1H, NH), 8.02 (m, 2H, H-5, H-8), 7.96 (m, 1H, H-6″), 7.85 (m, 3H, H-2″, H-6, H-7), 7.66 (m, 2H, H-4′, H-5″), 7.58 (d, J = 8.4 Hz, 1H, H-4″), 7.55 (s, 1H, H-2′), 7.27 (t, J = 7.9 Hz, 1H, H-5′), 7.03 (d, J = 7.9 Hz, 1H, H-6′), 4.00 (s, 2H, H-9), 2.19 (s, 3H, CH3) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 184.6 (C-4), 183.9 (C-1), 163.8 (CO(NH)), 148.3 (q, 3JC,F = 1.7 Hz, C-3″), 144.3 (C-2, C-3), 138.9 (C-3′), 138.6 (C-1′), 133.9 (C-6, C-7), 131.6 * (C-4a), 131.4 * (C-8a), 130.9 (C-1″), 130.5 (C-5″), 128.7 (C-5′), 126.7 (C-6″), 126.0 * (C-5), 125.9 * (C-8), 124.1 (C-6′), 123.9 (C-4″), 120.0 (C-2′, C-2″), 120.0 (q, 1JC,F = 256.9 Hz, CF3), 118.4 (C-4′), 31.7 (C-9), 13.0 (CH3) ppm; HRMS (ESI) calcd. for C26H17F3NO4 [M − H]− = 464.1110; found: 464.1115.

3.1.4. General Synthetic Procedure for Benzamido–Naphthoquinones 5a,b and 6a,b (Route B)

AgNO3 (0.1 equiv.) and (NH4)2S2O8 (2 equiv.) were dissolved in water (4 mL), and a solution of 1,4-naphthoquinone (200 mg, 1 equiv.) and the respective benzamido-phenylacetic acid derivative (1.4 equiv.) in a 1:1 mixture of CH3CN/CH2Cl2 (4 mL) was quickly added. The two-phase mixture was heated to 80 °C and stirred until the TLC showed complete consumption of the starting material (3–4 h). After cooling to ambient temperature, the mixture was extracted three times with CH2Cl2 (30 mL). The combined organic layers were washed with H2O, dried over Na2SO4, and concentrated in vacuo to give a residue, which was purified by flash chromatography as detailed below.
N-{4-[(1,4-Dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}benzamide (5a). Compound 5a was obtained after stirring for 3 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Amber solid; yield: 59%; Rf = 0.58 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 190–192 °C. 1H NMR (400 MHz, CDCl3): δ = 9.95 (s, 1H, NH), 8.10 (m, 1H, H-8), 8.04 (m, 1H, H-5), 7.87 (m, 2H, H-2″, H-6″), 7.73 (m, 2H, H-6, H-7), 7.62 (d, J = 7.7 Hz, 2H, H-3′, H-5′), 7.54 (m, 1H, H-4″), 7.48 (m, 2H, H-3″, H-5″), 7.25 (d, J = 7.7 Hz, 2H, H-2′, H-6′), 6.63 (s, 1H, H-3), 3.89 (s, 2H, H-9) ppm; 13C NMR (100 MHz, CDCl3): δ = 185.1 (C-4), 185.0 (C-1), 165.7 (CO(NH)), 150.8 (C-2), 136.9 (C-4′), 135.6 (C-3), 134.9 (C-1″), 133.8 (C-6, C-7), 132.9 (C-1′), 132.2 (C-8a), 132.1 (C-4a), 131.9 (C-4’′), 130.1 (C-2′, C-6′), 128.8 (C-3″, C-5″), 127.0 (C-2″, C-6″), 126.7 (C-8), 126.1 (C-5), 120.6 (C-3′, C-5′), 35.3 (C-9) ppm; HRMS (ESI) calcd. for C24H18NO3 [M + H]+ = 368.1287; found: 368.1277.
N-{4-[(1,4-Dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-4-(trifluoromethyl)benzamide (5b). Compound 5b was obtained after stirring for 4 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Amber solid; yield: 54%; Rf = 0.50 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 201–202 °C. 1H NMR (400 MHz, DMSO-d6): δ = 10.45 (s, 1H, NH), 8.13 (d, J = 8.1 Hz, 2H, H-2″, H-6″), 8.02 (m, 1H, H-8), 7.96 (m, 1H, H-5), 7.87 (m, 4H, H-6, H-7, H-3″, H-5″), 7.73 (d, J = 8.6 Hz, 2H, H-3′, H-5′), 7.30 (d, J = 8.6 Hz, 2H, H-2′, H-6′), 6.75 (s, 1H, H-3), 3.86 (s, 2H, H-9) ppm; 13C NMR (100 MHz, DMSO-d6): δ = 185.2 (C-4), 184.9 (C-1), 164.8 (CO(NH)), 151.0 (C-2), 139.2 (C-1″), 137.8 (C-4′), 135.7 (C-3), 134.6 (C-6, C-7), 133.5 (C-1′), 131.8 (q, 2JC,F = 31.9 Hz, C-4″) 132.2 (C-8a), 131.1 (C-4a), 130.6 (C-2″, C-6″), 130.0 (C-2′, C-6′), 126.7 (C-8), 126.1 (C-5), 125.8 (q, 3JC,F = 3.8 Hz, C-3″, C-5″), 124.3 (q, 1JC,F = 272.5 Hz, CF3), 121.1 (C-3′, C-5′), 34.9 (C-9) ppm; HRMS (ESI) calcd. for C25H15F3NO3 [M − H]− = 434.1004; found: 434.1012.
N-{3-[(1,4-Dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}benzamide (6a). Compound 6a was obtained after stirring for 3 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Amber solid; yield: 55%; Rf = 0.58 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 184–186 °C. 1H NMR (400 MHz, CDCl3): δ = 10.22 (s, 1H, NH), 8.01 (m, 1H, H-8), 7.97 (m, 1H, H-5), 7.92 (m, 2H, H-2″, H-6″), 7.86 (m, 2H, H-6, H-7), 7.70 (d, J = 8.1 Hz, 1H, H-4′), 7.68 (s, 1H, H-2′), 7.57 (m, 1H, H-4″), 7.51 (m, 2H, H-3″, H-5″), 7.31 (t, J = 8.1 Hz, 1H, H-5′), 7.06 (d, J = 7.6 Hz, 1H, H-6′), 6.81 (s, 1H, H-3), 3.88 (s, 2H, H-9) ppm; 13C NMR (100 MHz, CDCl3): δ = 184.6 (C-4), 184.3 (C-1), 165.5 (CO(NH)), 150.2 (C-2), 139.3 (C-3′), 137.9 (C-1′), 135.4 (C-3), 134.8 (C-1″), 134.1 (C-6, C-7), 131.7 * (C-8a), 131.5 * (C-4a), 131.5 (C-4″), 128.8 (C-5′), 128.3 (C-3″, C-5″), 127.6 (C-2″, C-6″), 126.2 (C-8), 125.6 (C-5), 124.5 (C-6′), 120.8 (C-2′), 118.6 (C-4′), 35.0 (C-9) ppm; HRMS (ESI) calcd. for C24H18NO3 [M + H]+ = 368.1287; found: 368.1276.
N-{3-[(1,4-Dioxo-1,4-dihydronaphthalen-2-yl)methyl]phenyl}-4-(trifluoromethyl)benzamide (6b). Compound 6b was obtained after stirring for 4 h, then purified by flash chromatography using cyclohexane:CH2Cl2:EtOAc (5:3:1). Amber solid; yield: 53%; Rf = 0.50 (cyclohexane:CH2Cl2:EtOAc = 5:3:1); m.p.: 205–206 °C. 1H NMR (400 MHz, CDCl3): δ = 8.08 (m, 1H, H-8), 8.02 (m, 1H, H-5), 7.96 (d, J = 7.8 Hz, 2H, H-2″, H-6″), 7.72 (m, 2H, H-6, H-7), 7.71 (d, J = 8.0 Hz, 2H, H-3″, H-5″), 7.57 (m, 1H, H-2′), 7.56 (d, J = 7.7 Hz, 1H, H-4′), 7.33 (t, J = 7.7 Hz, 1H, H-5′), 7.06 (dt, J = 7.7 Hz, 1.4 Hz, 1H, H-6′), 6.63 (t, J = 1.5 Hz, 1H, H-3), 3.89 (d, J = 1.5 Hz, 2H, H-9) ppm; 13C NMR (100 MHz, CDCl3): δ = 185.1 (C-4), 184.9 (C-1), 164.5 (CO(NH)), 150.4 (C-2), 138.1 (q, 5JC,F = 1.2 Hz, C-1″), 138.0 (C-1′, C-3′), 135.7 (C-3), 133.9 * (C-6), 133.8 * (C-7), 132.1 * (C-4a), 132.0 * (C-8a), 129.7 (C-5′), 127.6 (C-2″, C-6″), 126.7 (C-8), 126.1 (C-5), 126.0 (C-6′), 125.8 (q, 3JC,F = 3.9 Hz, C-3″, C-5″), 123.6 (q, 1JC,F = 272.6 Hz, CF3), 121.2 (C-2′), 119.0 (C-4′), 35.7 (C-9) ppm; HRMS (ESI) calcd. for C25H17F3NO3 [M + H]+ = 436.1161; found: 436.1146.

4. Conclusions

The present study describes the effective synthesis of 21 plasmodione-like benzamido–naphthoquinone derivatives, which exhibit remarkable antiplasmodial activity and selectivity. Utilizing advanced ligand efficiency metrics and structure–activity relationship analyses, we have identified highly effective and low-toxicity compounds, including the 2-fluoro-5-trifluoromethylbenzamido derivative 2f or the 2,4,5-trifluorobenzamido derivative 2i. The trypanocidal activity of the synthesized compounds was also evaluated. The strongest activity was observed in benzamido-NQ 5a, although both activity and selectivity were significantly less pronounced.
These findings highlight a promising pathway for developing new antiplasmodial drugs with superior efficacy and safety profiles. They also provide a robust framework for optimizing the drug discovery processes through the systematic application of physicochemical parameters.

Supplementary Materials

The following supporting information can be downloaded at https://www.mdpi.com/article/10.3390/ijms262210951/s1.

Author Contributions

Conceptualization, A.P.; methodology, A.P.; software, A.P.; validation, A.P. and W.S.; formal analysis, A.P.; investigation, A.P., G.B., E.-M.P.-W., M.C. and P.M.; resources, A.P., E.-M.P.-W., M.C. and P.M.; data curation, A.P., E.-M.P.-W., M.C., P.M. and W.S.; writing—original draft preparation, A.P. and W.S.; writing—review and editing, A.P., E.-M.P.-W., M.C., P.M. and W.S.; visualization, A.P.; supervision, A.P.; project administration, A.P.; funding acquisition, A.P. All authors have read and agreed to the published version of the manuscript.

Funding

Open Access Funding by the University of Graz.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The data presented in this study are available on request from the corresponding author.

Acknowledgments

The authors are grateful to R. Weis for discussions and to the University of Graz for open access funding. The authors gratefully acknowledge support from NAWI Graz for the Central Lab Environmental, Plant, and Microbial Metabolomics.

Conflicts of Interest

The authors declare no conflicts of interest.

Abbreviations

The following abbreviations are used in this manuscript:
AB-MPSAbbVie’s multi-parameter score
BEIBinding efficiency index
FQFit quality
H2LHit-to-lead
LELigand efficiency
LELPLigand efficiency lipophilicity price
LipELipophilic efficiency
LLELipophilic ligand efficiency
LLEATAstex lipophilic ligand efficiency
MDMenadione
MWMolecular weight
nBEINormalized binding efficiency index
NHANon-hydrogen atoms
NPOLNumber of oxygens and nitrogens in the structure
NQNaphthoquinone
NSEINormalized (polar) surface efficiency index
PFIGSK property forecast index
Ro5Rule of five
SEISurface efficiency index
SFISolubility forecast index
SILESize-independent ligand efficiency
TLCThin-layer chromatography
tPSATopological polar surface area
WHOWorld Health Organization

References

  1. Venkatesan, P. WHO world malaria report 2024. Lancet Microbe 2025, 6, 101073. [Google Scholar] [CrossRef]
  2. WHO Global Malaria Programme. World Malaria Report 2024. Available online: https://www.who.int/teams/global-malaria-programme/reports/world-malaria-report-2024 (accessed on 19 September 2025).
  3. Padhye, S.; Dandawate, P.; Yusufi, M.; Ahmad, A.; Sarkar, F.H. Perspectives on medicinal properties of plumbagin and its analogs. Med. Res. Rev. 2012, 32, 1131–1158. [Google Scholar] [CrossRef]
  4. Majiene, D.; Kuseliauskyte, J.; Stimbirys, A.; Jekabsone, A. Comparison of the effect of native 1,4-naphthoquinones plumbagin, menadione, and lawsone on viability, redox status, and mitochondrial functions of C6 glioblastoma cells. Nutrients 2019, 11, 1294. [Google Scholar] [CrossRef]
  5. de Souza, A.S.; Ribeiro, R.C.B.; Costa, D.C.S.; Pauli, F.P.; Pinho, D.R.; de Moraes, M.G.; Silva, F.d.C.d.; Forezi, L.d.S.M.; Ferreira, V.F. Menadione: A platform and a target to valuable compounds synthesis. Beilstein J. Org. Chem. 2022, 18, 381–419. [Google Scholar] [CrossRef] [PubMed]
  6. Dupouy, B.; Donzel, M.; Roignant, M.; Charital, S.; Keumoe, R.; Yamaryo-Botte, Y.; Feckler, A.; Bundschuh, M.; Bordat, Y.; Rottmann, M.; et al. 3-Benzylmenadiones and their Heteroaromatic Analogs Target the Apicoplast of Apicomplexa Parasites: Synthesis and Bioimaging Studies. ACS Infect. Dis. 2024, 10, 3553–3576. [Google Scholar] [CrossRef] [PubMed]
  7. Trometer, N.; Pecourneau, J.; Feng, L.; Navarro-Huerta, J.A.; Lazarin-Bidoia, D.; Lautenschlager, S.d.O.S.; Maes, L.; Francisco, A.F.; Kelly, J.M.; Meunier, B.; et al. Synthesis and Anti-Chagas Activity Profile of a Redox-Active Lead 3-Benzylmenadione Revealed by High-Content Imaging. ACS Infect. Dis. 2024, 10, 1808–1838. [Google Scholar] [CrossRef] [PubMed]
  8. Ehrhardt, K.; Deregnaucourt, C.; Goetz, A.-A.; Tzanova, T.; Gallo, V.; Arese, P.; Pradines, B.; Adjalley, S.H.; Bagrel, D.; Blandin, S.; et al. The redox cycler plasmodione is a fast-acting antimalarial lead compound with pronounced activity against sexual and early asexual blood-stage parasites. Antimicrob. Agents Chemother. 2016, 60, 5146–5158. [Google Scholar] [CrossRef]
  9. Sidorov, P.; Desta, I.; Chesse, M.; Horvath, D.; Marcou, G.; Varnek, A.; Davioud-Charvet, E.; Elhabiri, M. Redox Polypharmacology as an Emerging Strategy to Combat Malarial Parasites. ChemMedChem 2016, 11, 1339–1351. [Google Scholar] [CrossRef]
  10. Mounkoro, P.; Michel, T.; Golinelli-Cohen, M.-P.; Blandin, S.; Davioud-Charvet, E.; Meunier, B. A role for the succinate dehydrogenase in the mode of action of the redox-active antimalarial drug, plasmodione. Free Radic. Biol. Med. 2021, 162, 533–541. [Google Scholar] [CrossRef]
  11. Iacobucci, I.; Monaco, V.; Hovasse, A.; Dupouy, B.; Keumoe, R.; Cichocki, B.; Elhabiri, M.; Meunier, B.; Strub, J.-M.; Monti, M.; et al. Proteomic Profiling of Antimalarial Plasmodione Using 3-Benz(o)ylmenadione Affinity-Based Probes. ChemBioChem 2024, 25, e202400187. [Google Scholar] [CrossRef]
  12. Pinto, A.V.; de Castro, S.L. The trypanocidal activity of naphthoquinones: A review. Molecules 2009, 14, 4570–4590. [Google Scholar] [CrossRef]
  13. Futuro, D.O.; Ferreira, V.F.; Ferreira, P.G.; Nicoletti, C.D.; Borba-Santos, L.P.; Rozental, S.; Silva, F.C.D. The Antifungal Activity of Naphthoquinones: An Integrative Review. An. Da Acad. Bras. De Ciências 2018, 90, 1187–1214. [Google Scholar] [CrossRef] [PubMed]
  14. Patel, O.P.S.; Beteck, R.M.; Legoabe, L.J. Antimalarial application of quinones: A recent update. Eur. J. Med. Chem. 2021, 210, 113084. [Google Scholar] [CrossRef] [PubMed]
  15. Cárdenas, L.S.P.; Soler, K.A.A.; González, D.L.N.; Núñez, W.E.R.; Cárdenas-Chaparro, A.; Duchowicz, P.R.; Castaño, J.A.G. In Silico Antiprotozoal Evaluation of 1,4-Naphthoquinone Derivatives against Chagas and Leishmaniasis Diseases Using QSAR, Molecular Docking, and ADME Approaches. Pharmaceuticals 2022, 15, 687. [Google Scholar] [CrossRef] [PubMed]
  16. Santos, T.B.; de Moraes, L.G.C.; Pacheco, P.A.F.; dos Santos, D.G.; Ribeiro, R.M.d.A.C.; Moreira, C.d.S.; da Rocha, D.R. Naphthoquinones as a Promising Class of Compounds for Facing the Challenge of Parkinson’s Disease. Pharmaceuticals 2023, 16, 1577. [Google Scholar] [CrossRef]
  17. Bielitza, M.; Belorgey, D.; Ehrhardt, K.; Johann, L.; Lanfranchi, D.A.; Gallo, V.; Schwarzer, E.; Mohring, F.; Jortzik, E.; Williams, D.L.; et al. Antimalarial NADPH-Consuming Redox-Cyclers As Superior Glucose-6-Phosphate Dehydrogenase Deficiency Copycats. Antioxid. Redox Signal. 2015, 22, 1337–1351. [Google Scholar] [CrossRef]
  18. Presser, A.; Blaser, G.; Pferschy-Wenzig, E.-M.; Kaiser, M.; Maeser, P.; Schuehly, W. Pharmacomodulation of the Redox-Active Lead Plasmodione: Synthesis of Substituted 2-Benzylnaphthoquinone Derivatives, Antiplasmodial Activities, and Physicochemical Properties. Int. J. Mol. Sci. 2025, 26, 2114. [Google Scholar] [CrossRef]
  19. Rodo, E.C.; Feng, L.; Jida, M.; Ehrhardt, K.; Bielitza, M.; Boilevin, J.; Lanzer, M.; Williams, D.L.; Lanfranchi, D.A.; Davioud-Charvet, E. A Platform of Regioselective Methodologies to Access Polysubstituted 2-Methyl-1,4-naphthoquinone Derivatives: Scope and Limitations. Eur. J. Org. Chem. 2016, 2016, 1982–1993. [Google Scholar] [CrossRef]
  20. Donzel, M.; Karabiyikli, D.; Cotos, L.; Elhabiri, M.; Davioud-Charvet, E. Direct C-H Radical Alkylation of 1,4-Quinones. Eur. J. Org. Chem. 2021, 2021, 3622–3633. [Google Scholar] [CrossRef]
  21. Naturale, G.; Lamblin, M.; Commandeur, C.; Felpin, F.-X.; Dessolin, J. Direct C-H Alkylation of Naphthoquinones with Amino Acids Through a Revisited Kochi-Anderson Radical Decarboxylation: Trends in Reactivity and Applications. Eur. J. Org. Chem. 2012, 2012, 5774–5788. [Google Scholar] [CrossRef]
  22. Billamboz, M.; Bailly, F.; Barreca, M.L.; de Luca, L.; Mouscadet, J.-F.; Calmels, C.; Andreola, M.-L.; Witvrouw, M.; Christ, F.; Debyser, Z.; et al. Design, Synthesis, and Biological Evaluation of a Series of 2-Hydroxyisoquinoline-1,3(2H,4H)-diones as Dual Inhibitors of Human Immunodeficiency Virus Type 1 Integrase and the Reverse Transcriptase RNase H Domain. J. Med. Chem. 2008, 51, 7717–7730. [Google Scholar] [CrossRef]
  23. Nwaka, S.; Ramirez, B.; Brun, R.; Maes, L.; Douglas, F.; Ridley, R. Advancing drug innovation for neglected diseases-criteria for lead progression. PLoS Negl. Trop. Dis. 2009, 3, e440. [Google Scholar] [CrossRef] [PubMed]
  24. Katsuno, K.; Burrows, J.N.; Duncan, K.; van Huijsduijnen, R.H.; Kaneko, T.; Kita, K.; Mowbray, C.E.; Schmatz, D.; Warner, P.; Slingsby, B.T. Hit and lead criteria in drug discovery for infectious diseases of the developing world. Nat. Rev. Drug Discov. 2015, 14, 751–758. [Google Scholar] [CrossRef]
  25. Samby, K.; Willis, P.A.; Burrows, J.N.; Laleu, B.; Webborn, P.J.H. Actives from MMV Open Access Boxes? A suggested way forward. PLoS Pathog. 2021, 17, e1009384. [Google Scholar] [CrossRef]
  26. Banerjee, P.; Kemmler, E.; Dunkel, M.; Preissner, R. ProTox 3.0: A webserver for the prediction of toxicity of chemicals. Nucleic Acids Res. 2024, 52, W513–W520. [Google Scholar] [CrossRef]
  27. Pereira, V.S.d.S.; de Oliveira, C.B.S.; Fumagalli, F.; Emery, F.d.S.; da Silva, N.B.; de Andrade-Neto, V.F. Cytotoxicity, hemolysis and in vivo acute toxicity of 2-hydroxy-3-anilino-1,4-naphthoquinone derivatives. Toxicol. Rep. 2016, 3, 756–762. [Google Scholar] [CrossRef]
  28. Kombo, D.C.; LaMarche, M.J. The Logic of Chemical Optimization. J. Med. Chem. 2025, 68, 11572–11585. [Google Scholar] [CrossRef]
  29. Quancard, J.; Bach, A.; Borsari, C.; Craft, R.; Gnamm, C.; Gueret, S.M.; Hartung, I.V.; Koolman, H.F.; Laufer, S.; Lepri, S.; et al. The European Federation for Medicinal Chemistry and Chemical Biology (EFMC) Best Practice Initiative: Hit to Lead. ChemMedChem 2025, 20, e202400931. [Google Scholar] [CrossRef] [PubMed]
  30. Rees, D.C.; Congreve, M.; Murray, C.W.; Carr, R. Fragment-based lead discovery. Nat. Rev. Drug Discov. 2004, 3, 660–672. [Google Scholar] [CrossRef]
  31. Shultz, M.D. Two Decades under the Influence of the Rule of Five and the Changing Properties of Approved Oral Drugs. J. Med. Chem. 2019, 62, 1701–1714. [Google Scholar] [CrossRef] [PubMed]
  32. Tinworth, C.P.; Young, R.J. Facts, Patterns, and Principles in Drug Discovery: Appraising the Rule of 5 with Measured Physicochemical Data. J. Med. Chem. 2020, 63, 10091–10108. [Google Scholar] [CrossRef]
  33. Beckers, M.; Sturm, N.; Sirockin, F.; Fechner, N.; Stiefl, N. Prediction of Small-Molecule Developability Using Large-Scale In Silico ADMET Models. J. Med. Chem. 2023, 66, 14047–14060. [Google Scholar] [CrossRef]
  34. Davis, A.M.; Leeson, P.D. Physicochemical Properties. In The Handbook of Medicinal Chemistry: Principles and Practice; Ward, S.E., Davis, A., Eds.; The Royal Society of Chemistry: London, UK, 2023; pp. 1–39. ISBN 978-1-78801-898-2. [Google Scholar]
  35. Darlami, J.; Sharma, S. The role of physicochemical and topological parameters in drug design. Front. Drug Discov. 2024, 4, 1424402. [Google Scholar] [CrossRef]
  36. Meanwell, N.A. Improving Drug Design: An Update on Recent Applications of Efficiency Metrics, Strategies for Replacing Problematic Elements, and Compounds in Nontraditional Drug Space. Chem. Res. Toxicol. 2016, 29, 564–616. [Google Scholar] [CrossRef] [PubMed]
  37. Johnson, T.W.; Gallego, R.A.; Edwards, M.P. Lipophilic Efficiency as an Important Metric in Drug Design. J. Med. Chem. 2018, 61, 6401–6420. [Google Scholar] [CrossRef]
  38. Leeson, P.D.; Bento, A.P.; Gaulton, A.; Hersey, A.; Manners, E.J.; Radoux, C.J.; Leach, A.R. Target-Based Evaluation of “Drug-Like” Properties and Ligand Efficiencies. J. Med. Chem. 2021, 64, 7210–7230. [Google Scholar] [CrossRef]
  39. Nissink, J.W.M. Simple Size-Independent Measure of Ligand Efficiency. J. Chem. Inf. Model. 2009, 49, 1617–1622. [Google Scholar] [CrossRef] [PubMed]
  40. Reynolds, C.H.; Tounge, B.A.; Bembenek, S.D. Ligand binding efficiency: Trends, physical basis, and implications. J. Med. Chem. 2008, 51, 2432–2438. [Google Scholar] [CrossRef]
  41. Lipinski, C.A.; Lombardo, F.; Dominy, B.W.; Feeney, P.J. Experimental and computational approaches to estimate solubility and permeability in drug discovery and development settings. Adv. Drug Deliv. Rev. 2012, 64, 4–17. [Google Scholar] [CrossRef]
  42. DeGoey, D.A.; Chen, H.-J.; Cox, P.B.; Wendt, M.D. Beyond the Rule of 5: Lessons Learned from AbbVie’s Drugs and Compound Collection. J. Med. Chem. 2018, 61, 2636–2651. [Google Scholar] [CrossRef]
  43. Young, R.J.; Green, D.V.S.; Luscombe, C.N.; Hill, A.P. Getting physical in drug discovery II: The impact of chromatographic hydrophobicity measurements and aromaticity. Drug Discov. Today 2011, 16, 822–830. [Google Scholar] [CrossRef]
  44. McKerrow, J.H.; Lipinski, C.A. The rule of five should not impede anti-parasitic drug development. Int. J. Parasitol. Drugs Drug Resist. 2017, 7, 248–249. [Google Scholar] [CrossRef] [PubMed]
  45. Hopkins, A.L.; Keserue, G.M.; Leeson, P.D.; Rees, D.C.; Reynolds, C.H. The role of ligand efficiency metrics in drug discovery. Nat. Rev. Drug Discov. 2014, 13, 105–121. [Google Scholar] [CrossRef]
  46. Mortenson, P.N.; Murray, C.W. Assessing the lipophilicity of fragments and early hits. J. Comput.-Aided Mol. Des. 2011, 25, 663–667. [Google Scholar] [CrossRef] [PubMed]
  47. Abad-Zapatero, C.; Metz, J.T. Ligand efficiency indices as guideposts for drug discovery. Drug Discov. Today 2005, 10, 464–469. [Google Scholar] [CrossRef]
  48. Abad-Zapatero, C.; Blasi, D. Ligand Efficiency Indices (LEIs): More than a Simple Efficiency Yardstick. Mol. Inform. 2011, 30, 122–132. [Google Scholar] [CrossRef] [PubMed]
  49. Abad-Zapatero, C.; Champness, E.J.; Segall, M.D. Alternative variables in drug discovery: Promises and challenges. Future Med. Chem. 2014, 6, 577–593. [Google Scholar] [CrossRef]
  50. Abad-Zapatero, C. Ligand efficiency indices for effective drug discovery: A unifying vector formulation. Expert Opin. Drug Discov. 2021, 16, 763–775. [Google Scholar] [CrossRef]
Ijms 26 10951 g001
Figure 1. Alkyl-1,4-naphthoquinones with remarkable biological activities.
Figure 1. Alkyl-1,4-naphthoquinones with remarkable biological activities.
Ijms 26 10951 g001
Ijms 26 10951 sch001
Scheme 1. Synthesis of benzamido–menadione and naphthoquinone derivatives via Kochi–Anderson reaction. Reagents and conditions: (I) aminophenylacetic acid (1 equiv.), benzoyl chloride derivative (1.1 equiv.), 2 M KOH, rt, 18–24 h; (IIA) menadione (1 equiv.), phenylacetic acid derivative (1.4 equiv.), AgNO3 (0.35 equiv.), (NH4)2S2O8 (1.3 equiv.), CH3CN/H2O, 85 °C, 3–6 h; (IIB) 1,4-naphthoquinone (1 equiv.), phenylacetic acid derivative (1.4 equiv.), AgNO3 (0.1 equiv.), (NH4)2S2O8 (2 equiv.), CH3CN/CH2Cl2/H2O, 80 °C, 3–4 h.
Scheme 1. Synthesis of benzamido–menadione and naphthoquinone derivatives via Kochi–Anderson reaction. Reagents and conditions: (I) aminophenylacetic acid (1 equiv.), benzoyl chloride derivative (1.1 equiv.), 2 M KOH, rt, 18–24 h; (IIA) menadione (1 equiv.), phenylacetic acid derivative (1.4 equiv.), AgNO3 (0.35 equiv.), (NH4)2S2O8 (1.3 equiv.), CH3CN/H2O, 85 °C, 3–6 h; (IIB) 1,4-naphthoquinone (1 equiv.), phenylacetic acid derivative (1.4 equiv.), AgNO3 (0.1 equiv.), (NH4)2S2O8 (2 equiv.), CH3CN/CH2Cl2/H2O, 80 °C, 3–4 h.
Ijms 26 10951 sch001
Ijms 26 10951 g002
Figure 2. The scatterplot illustrates the relationship between lipophilic ligand efficiency (LLE) and size-independent ligand efficiency (SILE) of the synthesized compounds. This analysis also includes some optimization trajectories. The colours correspond to their respective pIC50 values, while the marker shapes correspond to the respective scaffolds.
Figure 2. The scatterplot illustrates the relationship between lipophilic ligand efficiency (LLE) and size-independent ligand efficiency (SILE) of the synthesized compounds. This analysis also includes some optimization trajectories. The colours correspond to their respective pIC50 values, while the marker shapes correspond to the respective scaffolds.
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Ijms 26 10951 g003
Figure 3. Plot of LLE (P. falciparum) data obtained in this study (x) vs. calculated log SI (P. falciparum) (y). The colours are assigned according to their respective pIC50 values, while the marker shapes are assigned to the respective scaffolds. Substances that lie in the yellow-shaded area combined high selectivity with acceptable ligand efficiency values.
Figure 3. Plot of LLE (P. falciparum) data obtained in this study (x) vs. calculated log SI (P. falciparum) (y). The colours are assigned according to their respective pIC50 values, while the marker shapes are assigned to the respective scaffolds. Substances that lie in the yellow-shaded area combined high selectivity with acceptable ligand efficiency values.
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Ijms 26 10951 g004
Figure 4. Application of the AtlasCBS concept, utilizing NSEI-nBEI plane analysis, to our synthesized compounds, encompassing the optimization of trajectories, with colours assigned according to the respective pIC50 values and marker shapes assigned to the respective scaffolds.
Figure 4. Application of the AtlasCBS concept, utilizing NSEI-nBEI plane analysis, to our synthesized compounds, encompassing the optimization of trajectories, with colours assigned according to the respective pIC50 values and marker shapes assigned to the respective scaffolds.
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Table 1. Synthesis of substituted benzamido–menadione and naphthoquinone derivatives (overall yields calculated from MD and NQ).
Table 1. Synthesis of substituted benzamido–menadione and naphthoquinone derivatives (overall yields calculated from MD and NQ).
CompdStructureRouteYield (%)CompdStructureRouteYield (%)
2aIjms 26 10951 i001A793cIjms 26 10951 i002A69
2bIjms 26 10951 i003A723dIjms 26 10951 i004A71
2cIjms 26 10951 i005A813eIjms 26 10951 i006A83
2dIjms 26 10951 i007A763fIjms 26 10951 i008A62
2eIjms 26 10951 i009A653gIjms 26 10951 i010A81
2fIjms 26 10951 i011A623hIjms 26 10951 i012A78
2gIjms 26 10951 i013A815aIjms 26 10951 i014B59
2hIjms 26 10951 i015A755bIjms 26 10951 i016B54
2iIjms 26 10951 i017A616aIjms 26 10951 i018B55
3aIjms 26 10951 i019A726bIjms 26 10951 i020B53
3bIjms 26 10951 i021A72
Table 2. In vitro antiparasitic activity, mammalian cell toxicity, and predicted oral toxicity of the synthesized compounds.
Table 2. In vitro antiparasitic activity, mammalian cell toxicity, and predicted oral toxicity of the synthesized compounds.
CompdP.falc. 1SI 2P.falc. 1T.b.rhod. 3SI 2T.b.rhod. 3Cyt L6 4Oral Toxicity 6
IC50 μM pIC50 5IC50 μM pIC50 5IC50 μMLD50 mg/kg
Chl.0.00910,0008.03 90750
Mel. 0.01512,0007.821803000
Pod. 0.019100
2a0.3364716.4764.10124.19158.174000
2b0.13616396.8734.58264.46222.511000
2c0.04547627.35160.62713.79213.941000
2d0.6633776.1884.44234.07250.364000
2e0.05329147.278.911175.05155.881000
2f0.02110,0007.67163.53713.79213.941000
2g0.02655807.5932.32444.49143.264000
2h0.06716857.18129.63613.89112.2257
2i0.02310,0007.64171.79613.76229.671600
3a1.487885.8365.33324.18130.954000
3b0.2654536.5827.68944.56119.824000
3c0.1675256.7865.34414.1887.574000
3d3.252695.4953.62844.27225.334000
3e1.0122115.9950.38344.30213.944000
3f0.4494766.3543.19554.36213.944000
3g14.46224.8426.06914.5830.551000
3h2.810765.5594.67924.02214.864000
5a4.55665.341.900145.7225.914000
5b14.97564.8215.01664.8283.621000
6a6.19235.213.50345.4615.594000
6b2.10475.685.23435.2814.054000
1 P. falciparum, strain NF54, erythrocytic stages; 2 SI is defined as the following ratio: IC50 in L6 cells/IC50 in each parasite; 3 T. brucei rhodesiense, strain STIB900 trypomastigote bloodstream forms; 4 cytotoxicity L6 cells rat skeletal myoblasts; 5 pIC50 = −log10 IC50 (M); 6 the oral toxicity was calculated using the ProTox 3.0 software (https://tox.charite.de/protox3/ (accessed on 4 July 2025)), and the calculated median lethal dose (LD50) is given in mg/kg body weight. Reference drugs: chloroquine (chl.), melarsoprol (mel.), and podophyllotoxin (pod.). Values in bold highlight the most striking results.
Table 3. Important ligand efficiency metrics and composite physicochemical descriptors, as well as their corresponding algebraic expressions and descriptions.
Table 3. Important ligand efficiency metrics and composite physicochemical descriptors, as well as their corresponding algebraic expressions and descriptions.
Variable NameDefinition
LE1.37 × (pIC50/NHA)
SILEpIC50/NHA0.3
FQ(pIC50/NHA)/[0.0715 + (7.5328/NHA) + (25.7079/NHA2) − (361.4722/NHA3)]
BEI(pIC50 × 1000)/MW
SEI(pIC50 × 100 Å2)/tPSA
LLE (or LipE)pIC50 − logP (or logD7.4)
LLEAT0.11 + [(1.37 × LLE)/NHA] = 0.11 + [(LE × LLE)/pIC50]
LELPlogP/LE
nBEIpIC50 + log10NHA
NSEIpIC50/NPOL(N+O)
PFI (SFI)logD7.4 + #aromrings
AB-MPS|logD7.4 − 3| + #rotbonds + #aromrings
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Presser, A.; Blaser, G.; Pferschy-Wenzig, E.-M.; Cal, M.; Mäser, P.; Schuehly, W. Synthesis, Antiprotozoal Activity, and Physicochemical Evaluation of Benzamido–Menadione Derivatives. Int. J. Mol. Sci. 2025, 26, 10951. https://doi.org/10.3390/ijms262210951

AMA Style

Presser A, Blaser G, Pferschy-Wenzig E-M, Cal M, Mäser P, Schuehly W. Synthesis, Antiprotozoal Activity, and Physicochemical Evaluation of Benzamido–Menadione Derivatives. International Journal of Molecular Sciences. 2025; 26(22):10951. https://doi.org/10.3390/ijms262210951

Chicago/Turabian Style

Presser, Armin, Gregor Blaser, Eva-Maria Pferschy-Wenzig, Monica Cal, Pascal Mäser, and Wolfgang Schuehly. 2025. "Synthesis, Antiprotozoal Activity, and Physicochemical Evaluation of Benzamido–Menadione Derivatives" International Journal of Molecular Sciences 26, no. 22: 10951. https://doi.org/10.3390/ijms262210951

APA Style

Presser, A., Blaser, G., Pferschy-Wenzig, E.-M., Cal, M., Mäser, P., & Schuehly, W. (2025). Synthesis, Antiprotozoal Activity, and Physicochemical Evaluation of Benzamido–Menadione Derivatives. International Journal of Molecular Sciences, 26(22), 10951. https://doi.org/10.3390/ijms262210951

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