Description:Background of Invention:
Antimicrobial resistance (AMR) has emerged as a critical concern with broad implications for our ability to combat infectious diseases caused primarily by microorganisms, including viruses, bacteria, fungi, and parasites [1-3]. Ongoing exposure to antibiotics has led to the evolution of multidrug-resistant (MDR) "superbugs" among pathogenic bacteria, posing severe threats to public health. In some cases, treatment options are severely limited, with heavy reliance on one or two specific medications. This situation raises a significant risk of pathogen resistance development, such as in the case of mucormycosis or black fungus infection, where the primary reliance is on Amphotericin B alone or a combination of Amphotericin B and Echinocandins [4]. Unfortunately, newly discovered antimicrobial drugs often exhibit cross-resistance and cannot effectively combat AMR due to their structural and mechanistic similarities to existing antibiotics [5,6]. Furthermore, the process of developing new drugs carries a substantial financial burden and is time-consuming, making them less accessible to underserved populations in regions where infectious diseases are prevalent [7].
To address these challenges, the development of novel chemical frameworks through efficient and rapid synthetic methods that produce diverse compounds without requiring a lengthy sequence of reactions can significantly accelerate the discovery of cost-effective drugs. In this context, the use of multicomponent reactions (MCRs) has gained significant attention for their ability to generate a wide range of compounds by sequentially combining three or more starting materials in one-pot reaction [8]. One such reaction is the Passerini reaction, discovered in Florence by Mario Passerini in 1921, which involves the reaction of isocyanides, aldehydes/ketones, and carboxylic acids to produce a-acyloxy carboxamides, representing the oldest isocyanide-based multicomponent reaction [9].
Long-chain fatty acids are widely recognized for their effectiveness in combating microbial pathogens. The antimicrobial properties of long alkyl chain fatty acids are influenced by the length of the carbon chain, which can be categorized into three primary groups: short (containing fewer than 8 carbon atoms), medium (ranging from 8 to 12 carbon atoms), and long (consisting of more than 12 carbon atoms). Additionally, the structure and efficacy of unsaturated fatty acids are affected by the number, location, and configuration (cis or trans) of their double bonds. In line with previous findings, it has been established that medium-chain saturated fatty acids and long-chain unsaturated fatty acids exhibit the highest level of activity against bacterial pathogens. Notably, gram-positive species display greater susceptibility to long-chain fatty acids when compared to gram-negative species. Moreover, in the case of long-chain unsaturated fatty acids, their antimicrobial potency increases with both the length of the carbon chain and the degree of unsaturation [10,11]]. Therefore, the design and synthesis of diverse alkyl chain-containing a-acyloxy carboxamides under mild reaction conditions represent a substantial and challenging objective. As depicted in Scheme 1, the Passerini reaction combines three components in a single step: an aldehyde, an acid, and an isocyanide to produce a-acyloxy amides. It is worth noting that this reaction employs straightforward reaction conditions, operates without the need for a catalyst, achieves 100% atom efficiency, highlighting its resource-efficient nature, and typically in good yields. These compounds hold applications span a variety of fields, including drug discovery, materials science, and chemical biology. In drug development, Passerini reactions play a pivotal role by enabling the rapid generation of new therapeutic compounds, expediting the search for novel antibiotics, antifungals, and other pharmaceutical agents [12-15]. Furthermore, they contribute to the design of functional materials such as polymers and catalytic converters. The versatility and efficiency of Passerini multi-component reactions position them as a valuable tool for researchers addressing a broad spectrum of scientific and industrial challenges.

Scheme 1. General one-pot Passerini reaction

Detailed description of the invention:
We employed the three-component Passerini reaction with modifications to generate the desired compounds [16,17]. This investigation highlights the synthesis of twenty new a-acyloxy carboxamides and proposes their potential as candidates for antimicrobial agents. All the starting materials, reagents and solvents were obtained from commercial suppliers (Sigma-Aldrich and Central Drug House) and used directly without any purification. The reactions, unless otherwise noted were carried out in oven dried glassware under nitrogen atmosphere. The synthesis of a-acyloxy amides (4a-4t) is detailed in Table 1. Initially, a mixture comprising acid (1.2 mmol), aldehyde (1 mmol), and isocyanide (1 mmol) was stirred at 25 oC in dichloromethane (4 mL). After 24 hours, the solvent was evaporated under reduced pressure. The resulting crude product underwent purification through column chromatography on silica gel, utilizing hexane/EtOAc (8:2) as the eluent. The desired products were obtained in solid form with yields ranging from 80% to 92%. Characterization of the products was done by determining their melting points and analyzing their spectral data including IR, 1H NMR, 13C NMR, and HRMS. In the 1H NMR spectrum, a sharp singlet at d 6.00 confirms the presence of the -NH group due to the isocyanide core (4a). In the 13C NMR spectrum, signals at d 166.3 and 169.0 ppm are indicative of the carbonyl groups in the amide and ester moieties. The FTIR spectrum exhibits distinct peaks at 3285 cm-1, corresponding to NH bond stretching in the amide. Additionally, peaks at 2996 cm-1, 2969 cm-1, and 2931 cm-1 are associated with CH stretching in the alkyl chains, while peaks at 1746 cm-1 and 1654 cm-1 represent stretching vibrations of the carbonyl >C=O groups in the ester and amide functional groups.

After successfully characterizing the synthesized compounds, we conducted tests to evaluate their antimicrobial properties, both against bacteria and fungi. To assess the antimicrobial activity of these compounds, a cell susceptibility assay was conducted using gram-positive (S. aureus) and gram-negative (E. coli) bacteria, as well as a fungus (C. albicans). In this analysis, a fixed amount (60 µg) of the synthesized compounds was employed for a spot assay. The initial assessment of the antibacterial and antifungal effectiveness of these compounds was based on the observed zones of inhibition on the plates. Compounds showing promising results in these preliminary tests were then selected for further evaluation, including the determination of IC50 values through a broth microdilution assay.

Experimental Section
General method for synthesis of Passerini a-acyloxy amides
Initially, acid 1 (1.2 mmol) and aldehyde 2 (1 mmol) were dissolved in 4 mL of CH2Cl2 and stirred for 20 minutes at 25 oC. Subsequently, isocyanide 3 (1 mmol) was added dropwise to the reaction mixture. The resulting reaction mixture was stirred for 24 hours at 25 oC, and its progress was monitored by TLC. Solvents were evaporated under reduced pressure after completion of the reaction. Following this, 10 mL of CH2Cl2 and 10 mL of saturated NaHCO3 were added, and the mixture was extracted with CH2Cl2 (3 x 10 mL). The organic layer was then dried over Na2SO4, filtered, and the solvent was removed under vacuum. The resulting crude product underwent purification via silica-gel column chromatography and/or recrystallization.

Susceptibility assessment: The antibacterial and antifungal properties of the synthesized compounds were evaluated through spot and microdilution assays. In the spot assay, both gram-positive (S. aureus) and gram-negative (E. coli) bacteria [18,19], as well as the fungal pathogen C. albicans, were cultured in LB, MHB, and YEPD broth, respectively [20-22]. Cell cultures were grown until they reached a cell density of 0.5 at 600 nm, representing the mid-exponential phase, and the cells were then harvested to obtain the cell pellet. This pellet was resuspended in PBS to maintain an optical density (OD) of 0.1, and an equal number of cells were evenly spread on agar plates. A fixed quantity of 60 µg of each synthesized compound was spotted onto the plates. Subsequently, the plates were incubated (bacterial plates at 37 °C and fungal plates at 30 °C) overnight to observe the zones of inhibition. The effectiveness of the synthesized compounds was assessed based on the size of the inhibition zones, and those showing promise were selected for further IC50 value determination using the broth microdilution assay. In the broth microdilution assay, IC50 values were determined following established methods [23-25]. Different dilutions of the synthesized compounds were prepared in a 96-well plate, starting with the highest concentration of 10 mg/mL in the first well and further serially diluted by 2-fold increments until reaching a concentration of 0.156 mg/mL. After successful dilutions, a specific number of mid-exponential phase cells from S. aureus, E. coli, and C. albicans were added to each well to maintain a final volume of 200 µL in each well, and the plate was then incubated at varying temperatures. Bacterial cultures were incubated at 37 °C, and fungal cultures were incubated at 30 °C, each for 24 hours. The IC50 values were determined by assessing the optical density (OD600nm) of the incubated plate.

Results and Discussion
Characterization data of the compound 4a-t:
2-(tert-Butylamino)-1-(4-cyanophenyl)-2-oxoethyl acetate (4a)
Yield: 83%; Off white solid; m.p. = 134-136 °C; IR (neat): ?max = 3285 (NH str.), 2227 (C=N str.), 1746 (CO str. ester), 1654 (CO str. amide) cm-1; 1H NMR (400 MHz, CDCl3) d 7.63-7.57 (m, 2H), 7.53 (d, J = 8.2 Hz, 2H), 6.29 (s, 1H), 5.92 (d, J = 1.8 Hz, 1H), 2.21-2.12 (m, 3H), 1.34-1.24 (m, 9H); 13C NMR (101 MHz, CDCl3) d 168.9, 166.3, 141.2, 132.3, 127.9, 118.4, 112.3, 74.7, 51.7, 28.5, 20.9; HRMS calcd for C15H18N2O3 [M+1]+, 275.1391; found, 275.1396.
2-(tert-Butylamino)-1-(4-cyanophenyl)-2-oxoethyl propionate (4b)
Yield: 85%; Off white solid; m.p. = 120-122 °C; IR (neat): ?max = 3290 (NH str.), 2227 (C=N str.), 1742 (CO str. ester), 1650 (CO str. amide), 2227 (C=N str.) cm-1; 1H NMR (400 MHz, CDCl3) d 7.66 (d, J = 8.3 Hz, 2H), 7.54 (d, J = 8.2 Hz, 2H), 6.06 (s, 1H), 5.99 (s, 1H), 2.58-2.41 (m, 2H), 1.36 (s, 9H), 1.20 (t, J = 7.5 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 172.1, 166.2, 141.1, 132.4, 127.8, 118.4, 112.6, 74.5, 51.7, 29.7, 28.6, 27.5, 8.9; HRMS calcd for C16H20N2O3 [M+1]+, 289.1552; found, 289.1555
2-(tert-Butylamino)-1-(4-cyanophenyl)-2-oxoethyl 4-oxopentanoate (4c)
Yield: 83%; Off white solid; m.p. = 80-82 °C; IR (neat): ?max = 3292 (NH str.), 2227 (C=N str.), 1738 (CO str. ester), 1650 (CO str. amide) cm-1; 1H NMR (400 MHz, CDCl3) d 7.66-7.52 (m, 2H), 7.50 (d, J = 8.2 Hz, 2H), 6.52 (s, 1H), 5.91 (s, 1H), 2.84-2.73 (m, 2H), 2.71-2.62 (m, 1H), 2.52 (m, 1H), 2.11 (s, 3H), 1.28 (s, 9H); 13C NMR (101 MHz, CDCl3) d 207.0, 171.1, 166.5, 141.0, 132.2, 127.9, 118.4, 112.2, 74.8, 51.7, 38.0, 29.6, 28.4, 28.0; HRMS calcd for C18H22N2O4 [M+1]+, 331.1658; found, 331.1663.

2-(tert-Butylamino)-1-(4-cyanophenyl)-2-oxoethyl hexanoate (4d)
Yield: 84%; Off white solid; m.p. = 74-76 °C; IR (neat): ?max = 3290 (NH str.), 2227 (C=N str.), 1744 (CO str. ester), 1654 (CO str. amide) cm-1; 1H NMR (400 MHz, CDCl3) 1H NMR (400 MHz, CDCl3) d 7.65 (d, J = 8.2 Hz, 2H), 7.55 (d, J = 8.2 Hz, 2H), 6.15 (s, 1H), 5.99 (s, 1H), 2.46 (td, J = 7.5, 3.8 Hz, 2H), 1.65 (dd, J = 14.0, 7.0 Hz, 2H), 1.35 (s, 9H), 1.25 (s, 4H), 0.87 (t, J = 6.6 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 171.5, 166.3, 141.2, 132.3, 127.8, 118.4, 112.5, 74.4, 51.7, 34.1, 31.8, 29.1, 28.5, 22.6, 14.0; HRMS calcd for C19H26N2O3 [M+1]+, 331.2022; found, 331.2020.
2-(tert-Butylamino)-1-(4-cyanophenyl)-2-oxoethyl decanoate (4e)
Yield: 87%; Off white solid; m.p. = 76-78 °C; IR (neat): ?max = 3306 (NH str.), 2227 (C=N str.), 1742 (CO str. ester), 1658 (CO str. amide) cm-1; 1H NMR (400 MHz, CDCl3) 1H NMR (400 MHz, CDCl3) d 7.68 (d, J = 8.3 Hz, 2H), 7.56 (d, J = 8.2 Hz, 2H), 6.09 (s, 1H), 6.01 (s, 1H), 2.47 (td, J = 7.5, 3.4 Hz, 2H), 1.69 (dt, J = 14.4, 7.3 Hz, 4H), 1.46-1.24 (m, 19H), 0.91 (t, J = 6.9 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 171.5, 166.2, 141.1, 132.4, 127.8, 118.4, 112.5, 74.5, 51.7, 34.1, 31.1, 28.6, 24.5, 22.2, 13.8; HRMS calcd for C23H34N2O3 [M+1]+, 387.2648; found, 387.2648.
2-(tert-Butylamino)-1-(4-cyanophenyl)-2-oxoethyl dodecanoate (4f)
Yield: 89%; Off white solid; m.p. = 66-68 °C; IR (neat): ?max = 3294 (NH str.), 2227 (C=N str.), 1740 (CO str. ester), 1654 (CO str. amide) cm-1; 1H NMR (400 MHz, CDCl3) 1H NMR (400 MHz, CDCl3) d 7.67 (d, J = 8.2 Hz, 2H), 7.56 (d, J = 8.2 Hz, 2H), 6.12 (s, 1H), 6.01 (s, 1H), 2.47 (m, 2H), 2.35 (t, J = 7.5 Hz, 1H), 1.66 (m, 2H), 1.37 (s, 9H), 1.28 (d, J = 12.3 Hz, 15H), 0.89 (t, J = 6.8 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 171.5, 166.3, 141.1, 132.4, 127.8, 118.4, 112.5, 74.5, 51.7, 34.2, 33.9, 31.8, 29.6, 28.4, 29.3, 29.2, 29.0, 28.6, 28.6, 24.7, 22.6, 14.1; HRMS calcd for C25H38N2O3 [M+1]+, 415.2961; found, 415.2964.
2-(tert-Butylamino)-1-(4-cyanophenyl)-2-oxoethyl tetradecanoate (4g)
Yield: 88%; Off white solid; m.p. = 60-62 °C; IR (neat): ?max = 3305 (NH str.), 2227 (C=N str.), 1740 (CO str. ester), 1656 (CO str. amide) cm-1; 1H NMR (400 MHz, CDCl3) 1H NMR (400 MHz, CDCl3) d 7.65 (d, J = 8.0 Hz, 2H), 7.55 (d, J = 8.3 Hz, 2H), 6.19 (s, 1H), 6.00 (s, 1H), 2.51-2.40 (m, 2H), 2.35 (dd, J = 24.6, 17.0 Hz, 1H), 1.64 (dt, J = 14.5, 7.0 Hz, 3H), 1.35 (s, 9H), 1.25 (s, 18H), 0.87 (t, J = 6.5 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 178.9, 171.6, 166.4, 141.2, 132.4, 127.9, 118.4, 112.5, 74.4, 51.8, 31.9, 29.6, 29.4, 29.3, 29.2, 28.6, 22.7, 14.1; HRMS calcd for C27H42N2O3 [M+1]+, 443.3274; found, 443.3273.
2-(tert-Butylamino)-1-(4-cyanophenyl)-2-oxoethyl palmitate (4h)
Yield: 90%; Off white solid; m.p. = 79-81 °C; IR (neat): ?max = 3304 (NH str.), 2227 (C=N str.), 1744 (CO str. ester), 1656 (CO str. amide) cm-1; 1H NMR (400 MHz, CDCl3) 1H NMR (400 MHz, CDCl3) d 7.59 (d, J = 8.4 Hz, 2H), 7.47 (d, J = 8.2 Hz, 2H), 6.00 (s, 1H), 5.92 (s, 1H), 2.38 (td, J = 7.5, 3.4 Hz, 2H), 1.68-1.54 (m, 2H), 1.29 (s, 9H), 1.18 (s, 24H), 0.82 (d, J = 6.6 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 171.5, 166.3, 141.1, 132.4, 127.8, 118.4, 112.6, 74.5, 51.7, 34.2, 31.9, 29.6, 29.6, 29.6, 29.5, 29.4, 29.3, 29.2, 29.0, 28.6, 24.8, 22.7, 14.1; HRMS calcd for C29H46N2O3 [M+1]+, 471.3587; found, 471.3590.
2-(tert-Butylamino)-1-(4-cyanophenyl)-2-oxoethyl stearate (4i)
Yield: 86%; Off white solid; m.p. = 73-75 °C; IR (neat): ?max = 3294 (NH str.), 2227 (C=N str.), 1740 (CO str. ester), 1654 (CO str. amide) cm-1; 1H NMR (400 MHz, CDCl3) 1H NMR (400 MHz, CDCl3) d 7.71-7.64 (m, 2H), 7.61-7.49 (m, 2H), 6.19-6.07 (m, 1H), 6.02-5.97 (m, 1H), 2.46 (td, J = 7.5, 3.9 Hz, 2H), 1.66 (dd, J = 14.0, 7.0 Hz, 2H), 1.36 (d, J = 1.3 Hz, 9H), 1.26 (s, 28H), 0.88 (dd, J = 6.8, 5.9 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 171.6, 166.3, 141.1, 132.4, 127.8, 118.4, 112.5, 74.5, 51.7, 34.2, 31.0, 29.7, 29.6, 29.4, 28.6, 24.8, 22.7, 14.1; HRMS calcd for C31H50N2O3 [M+1]+, 499.3892; found, 499.3900.
2-(tert-Butylamino)-1-(4-cyanophenyl)-2-oxoethyl oleate (4j)
Yield: 88%; Off white solid; m.p. = 75-77 °C; IR (neat): ?max = 3307 (NH str.), 2227 (C=N str.), 1742 (CO str. ester), 1656 (CO str. amide) cm-1; 1H NMR (400 MHz, CDCl3) 1H NMR (400 MHz, CDCl3) d 7.67 (d, J = 8.2 Hz, 2H), 7.55 (d, J = 8.3 Hz, 2H), 6.13 (s, 1H), 6.00 (s, 1H), 5.35 (d, J = 5.0 Hz, 2H), 2.47 (td, J = 7.5, 4.0 Hz, 2H), 2.02 (d, J = 5.8 Hz, 2H), 1.70-1.61 (m, 2H), 1.36 (s, 9H), 1.29 (d, J = 14.6 Hz, 22H), 0.88 (t, J = 6.7 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 171.5, 166.3, 141.2, 132.4, 130.1, 130.0, 129.9, 129.7, 129.6, 128.2, 118.4, 112.6, 74.5, 51.7, 34.1, 31.9, 29.7, 29.5, 29.3, 29.0, 28.6, 27.1, 24.8, 22.6, 14.1; HRMS calcd for C31H48N2O3 [M+1]+, 497.3743; found, 497.3747.
1-(4-Cyanophenyl)-2-(cyclohexylamino)-2-oxoethyl stearate (4k)
Yield: 87%; Off white solid; m.p. = 80-83 °C; IR (neat): ?max = 3285 (NH str.), 2224 (C=N str.), 1742 (CO str. ester), 1654 (CO str. amide) cm-1; 1H NMR (400 MHz, CDCl3) 1H NMR (400 MHz, CDCl3) d 7.67 (d, J = 8.3 Hz, 2H), 7.57 (d, J = 8.3 Hz, 2H), 6.23-6.15 (m, 1H), 6.10 (s, 1H), 3.92-3.63 (m, 1H), 2.48 (d, J = 3.8 Hz, 2H), 1.91 (s, 3H), 1.67 (dd, J = 14.3, 7.1 Hz, 9H), 1.27 (s, 28H), 0.89 (s, 3H); 13C NMR (101 MHz, CDCl3) d 171.5, 166.3, 141.0, 132.4, 127.8, 118.4, 112.6, 74.3, 48.3, 34.0, 32.8, 31.9, 29.7, 29.6, 29.4, 29.3, 29.2, 29.2, 29.0, 29.0, 25.3, 22.7, 14.1; HRMS calcd for C33H52N2O3 [M+1]+, 525.4069; found, 525.4056.
2-(tert-Butylamino)-1-(4-nitrophenyl)-2-oxoethyl tetradecanoate (4l)
Yield: 92%; Off white solid; m.p. = 74-76 °C; IR (neat): ?max = 3271 (NH str.), 1735 (CO str. ester), 1660 (CO str. amide), 1344 and 1528 (NO str.) cm-1; 1H NMR (400 MHz, CDCl3) d 7.91 (d, J = 7.9 Hz, 1H), 7.69 (d, J = 7.4 Hz, 1H), 7.58 (t, J = 7.2 Hz, 1H), 7.43 (t, J = 7.3 Hz, 1H), 6.43 (s, 1H), 6.15 (s, 1H), 2.35 (dd, J = 13.5, 6.7 Hz, 2H), 1.57 (s, 3H), 1.27 (s, 9H), 1.18 (s, 19H), 0.80 (d, J = 5.9 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 172.4, 165.9, 148.0, 133.5, 131.0, 130.1, 129.4, 124.7, 71.3, 51.7, 34.0, 31.9, 29.6, 29.5, 29.4, 29.3, 29.2, 29.0, 28.5, 24.7, 22.6, 14.1; HRMS calcd for C26H42N2O5 [M+1]+, 463.3172; found, 463.3176.

2-(tert-Butylamino)-1-(4-nitrophenyl)-2-oxoethyl stearate (4m)
Yield: 89%; Off white solid; m.p. = 66-68 °C; IR (neat): ?max = 3290 (NH str.), 1744 (CO str. ester), 1652 (CO str. amide), 1348 and 1522 (NO str.) cm-1; 1H NMR (400 MHz, CDCl3) d 8.14 (d, J = 8.8 Hz, 2H), 7.54 (d, J = 8.6 Hz, 2H), 6.07 (s, 1H), 5.97 (s, 1H), 2.40 (td, J = 7.5, 3.5 Hz, 2H), 1.67-1.54 (m, 2H), 1.29 (d, J = 6.4 Hz, 10H), 1.20 (d, J = 18.5 Hz, 27H), 0.80 (t, J = 6.9 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 171.5, 166.2, 148.0, 143.0, 128.0, 123.8, 74.3, 51.8, 34.2, 31.9, 29.6, 28.6, 24.8, 22.7, 14.1; HRMS calcd for C30H50N2O5 [M+1]+, 519.3797; found, 519.3798.
2-(Cyclohexylamino)-1-(4-nitrophenyl)-2-oxoethyl stearate (4n)
Yield: 90%; Off white solid; m.p. = 78-80 °C; IR (neat): ?max = 3284 (NH str.), 1746 (CO str. ester), 1658 (CO str. amide), 1346 and 1521 (NO str.) cm-1; 1H NMR (400 MHz, CDCl3) d 8.23 (d, J = 8.5 Hz, 2H), 7.64 (d, J = 8.5 Hz, 2H), 6.15 (s, 2H), 3.89-3.66 (m, 1H), 2.49 (d, J = 3.6 Hz, 3H), 1.92 (s, 2H), 1.70 (dd, J = 16.4, 10.1 Hz, 6H), 1.29 (d, J = 18.1 Hz, 31H), 0.89 (t, J = 6.4 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 171.5, 166.1, 148.0, 142.9, 128.0, 123.7, 74.1, 48.4, 34.1, 32.8, 31.9, 29.7, 29.6, 29.6, 29.4, 29.3, 29.2, 29.0, 25.3, 24.7, 22.7, 14.1; HRMS calcd for C32H52N2O5 [M+1]+, 545.3954; found, 545.3958.
2-(tert-Butylamino)-1-(4-chlorophenyl)-2-oxoethyl stearate (4o)
Yield: 88%; Off white solid; m.p. = 68-71 °C; IR (neat): ?max = 3287 (NH str.), 1742 (CO str. ester), 1652 (CO str. amide), 722 (C-Cl str.) cm-1; 1H NMR (400 MHz, CDCl3) d 7.27 (d, J = 1.1 Hz, 4H), 5.91 (s, 1H), 5.86 (s, 1H), 2.35 (d, J = 3.4 Hz, 2H), 1.57 (d, J = 7.3 Hz, 5H), 1.29 (s, 9H), 1.18 (s, 25H), 0.80 (d, J = 7.0 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 179.1, 171.7, 167.1, 134.7, 134.7, 134.6, 128.8, 128.82, 128.7, 74.6, 51.6, 35.3, 34.2, 34.0, 31.9, 29.7, 29.6, 29.4, 28.6, 24.8, 22.7, 14.1; HRMS calcd for C30H50ClNO3 [M+1]+, 508.3557; found, 508.3553.

1-(4-Bromophenyl)-2-(tert-butylamino)-2-oxoethyl tetradecanoate (4p)
Yield: 85%; Off white solid; m.p. = 76-78 °C; IR (neat): ?max = 3282 (NH str.), 1742 (CO str. ester), 1652 (CO str. amide), 669 (C-Br str.) cm-1; 1H NMR (400 MHz, CDCl3) d 7.50-7.38 (m, 2H), 7.26 – 7.18 (m, 2H), 5.94 (s, 1H), 5.85 (s, 1H), 2.35 (td, J = 7.5, 3.4 Hz, 2H), 2.27 (d, J = 7.5 Hz, 1H), 1.63-1.51 (m, 3H), 1.28 (s, 9H), 1.18 (s, 18H), 0.80 (d, J = 7.0 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 179.0, 171.7, 167.0, 135.1, 131.8, 129.0, 122.9, 74.7, 51.6, 34.2, 33.9, 31.9, 29.7, 29.6, 29.5, 29.4, 29.3, 29.2, 28.6, 24.8, 22.6, 14.1; HRMS calcd for C26H42 BrNO3 [M+1]+, 496.2426; found, 496.2427.

1-(4-Bromophenyl)-2-(tert-butylamino)-2-oxoethyl stearate (4q)
Yield: 84%; Off off-white solid; m.p. = 72-74 °C; IR (neat): ?max = 3285 (NH str.), 1742 (CO str. ester), 1652 (CO str. amide), 654 (C-Br str.) cm-1; 1H NMR (400 MHz, CDCl3) d 7.49 (d, J = 8.5 Hz, 2H), 7.29 (d, J = 8.4 Hz, 2H), 5.96 (d, J = 26.5 Hz, 2H), 2.53 – 2.30 (m, 3H), 1.64 (dd, J = 14.5, 7.3 Hz, 3H), 1.35 (s, 9H), 1.26 (s, 26H), 0.87 (d, J = 7.0 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 178.9, 171.7, 167.0, 135.1, 131.8, 129.0, 122.9, 74.7, 51.6, 34.2, 33.9, 31.9, 29.7, 29.6, 29.5, 29.4, 29.3, 29.2, 29.0, 28.6, 24.8, 22.7, 14.1; HRMS calcd for C30H50BrNO3 [M+1]+, 552.3052; found, 552.3047.

2-(tert-Butylamino)-1-(4-fluorophenyl)-2-oxoethyl stearate (4r)
Yield: 86%; Off white solid; m.p. = 56-58 °C; IR (neat): ?max = 3290 (NH str.), 1742 (CO str. ester), 1654 (CO str. amide), 1159 (C-F str.) cm-1; 1H NMR (400 MHz, CDCl3) d 7.41 (dd, J = 8.1, 5.5 Hz, 2H), 7.04 (t, J = 8.5 Hz, 2H), 6.11 (s, 1H), 5.98 (s, 1H), 2.51-2.37 (m, 2H), 2.33 (t, J = 7.5 Hz, 1H), 1.64 (dd, J = 14.2, 7.1 Hz, 4H), 1.36 (s, 9H), 1.27 (s, 25H), 0.89 (d, J = 6.1 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 179.1, 171.8, 167.4, 164.1, 132.0, 129.3, 115.7, 115.4, 74.6, 51.5, 34.1, 31.9, 29.7, 29.6, 29.4, 29.2, 28.6, 24.9, 22.7, 14.1; 19F NMR (376 MHz, CDCl3) d -112.75 (m, 1F); HRMS calcd for C30H50FNO3 [M+1]+, 492.3853; found, 492.3857.

2-(tert-Butylamino)-2-oxo-1-(3,4,5-trimethoxyphenyl)ethyl tetradecanoate (4s)
Yield: 80%; Off white solid; m.p. = 52-54 °C; IR (neat): ?max = 3331 (NH str.), 1736 (CO str. ester), 1658 (CO str. amide), 1123 (C-O-C str.) cm-1; 1H NMR (400 MHz, CDCl3) d 6.63 (s, 2H), 5.98 (s, 1H), 5.92 (s, 1H), 3.85 (d, J = 9.3 Hz, 9H), 2.52-2.38 (m, 2H), 1.66 (dd, J = 14.5, 7.2 Hz, 2H), 1.38 (s, 9H), 1.25 (s, 20H), 0.88 (t, J = 6.5 Hz, 3H); 13C NMR (101 MHz, CDCl3) d 177.9, 172.0, 167.4, 153.3, 138.3, 131.5, 104.7, 75.3, 60.7, 56.1, 51.5, 34.3, 33.8, 31.9, 29.6, 29.3, 29.1, 28.7, 24.8, 22.6, 14.1.
2-(tert-Butylamino)-2-oxo-1-(pyridin-2-yl)ethyl stearate (4t)
Yield: 84%; Off white solid; m.p. = 48-50 °C; IR (neat): ?max = 3255 (NH str.), 1739 (CO str. ester), 1687 (CO str. amide), 1469 (C=N str.) cm-1; 1H NMR (400 MHz, CDCl3) d 8.60-8.49 (m, 1H), 7.79-7.64 (m, 1H), 7.52 (d, J = 7.7 Hz, 1H), 7.31-7.17 (m, 1H), 6.74 (d, J = 29.9 Hz, 1H), 6.02 (s, 1H), 2.56-2.41 (m, 2H), 1.77-1.56 (m, 2H), 1.34-1.28 (m, 12H), 1.23 (s, 25H), 0.89-0.82 (m, 3H); 13C NMR (101 MHz, CDCl3) d 172.1, 166.0, 155.3, 148.9, 137.1, 123.3, 122.1, 75.5, 51.4, 34.0, 31.9, 29.6, 29.5, 29.4, 29.3, 29.2, 29.0, 28.5, 24.7, 22.6, 14.1; HRMS calcd for C29H50N2O3 [M+1]+, 475.3900; found, 475.3909.
Anti-microbial assessment: The antibacterial effect of synthesized compounds was assessed using gram –ve E. coli and gram +ve S. aureus bacteria and the antifungal effect was measured using C. albicans. Our results showed that synthesized compound 4f hypersensitizes both E. coli and S. aureus with no significant inhibitory effect on C. albicans cells (Table 2). However, compounds; 4d, 4i, 4k and 4m elicited potent antifungal activity with lower IC50 values shown in Table 3. Of note, other tested compounds showed either no or mild antibacterial and antifungal effects.

Table 2: Antimicrobial screening of synthesized compounds
Compounds Activity against
E. coli Activity against
S. aureus Activity against
C. albicans
4a + + -
4b - - -
4c - + -
4d - - +++
4e - - ++
4f +++ +++ -
4g + +++ ++
4h - - ++
4i + - +++
4j + - -
4k + - +++
4l + - -
4m ++ - +++
4n - ++ -
4o - - -
4p - - -
4q - - -
4r - + -
4s ++ - ++
4t - - -

Table 3: IC50 values of the tested compounds
Compounds IC50 (mg/mL) value for E. coli IC50 (mg/mL) value for S. aureus IC50 (mg/mL) value for C. albicans
4a 1.8320 1.669 ND
4c ND 1.305 ND
4d ND ND 3.330
4e ND ND 4.240
4f 0.1140 0.7764 ND
4g 0.6220 0.9942 6.626
4h ND ND 4.289
4i 0.6345 ND 3.075
4j 0.9169 ND ND
4k 0.7594 ND 3.821
4l 0.3139 ND ND
4m 0.2057 ND 3.851
4n ND 1.289 ND
4r ND 1.152 ND
4s 0.2562 ND 5.710
ND: Not detectable
, Claims:Claims:
I/We claim:
1. A process for preparing a-acyloxy carboxamides via Passerini reaction comprising:
(i) Acid (1.2 mmol) and aldehyde (1 mmol) was dissolved in CH2Cl2 (4 mL) in a round-bottom flask of size 10 mL on a magnetic stirrer with 450 rpm for 20 minutes.
(ii) Then isocyanide (1 mmol) was added in a dropwise manner into the above mixture under constant stirring of 450 rpm.
(iii) The reaction mixture was left overnight at 25 °C under constant stirring of 450 rpm.
(iv) Then, progress of reaction was monitored with the help of TLC. After completion of the reaction, solvent was dried under reduced pressure with the help of rotatory evaporator.
(v) Afterwards, dichloromethane (10 mL) and saturated sodium bicarbonate (10 mL) was added and extracted with CH2Cl2 (3 x 10 mL).
(vi) After drying the organic layer over Na2SO4 (0.003 g), the solvent was removed in vacuo and the crude product was purified by column chromatography (EtOAc:Hexane; 2:8) and/or recrystallization.

2. The compounds synthesized from the process of Claim (1) with structural formula-(I) via Passerini three component reaction, procedure as claimed in claim 1, in which R is 4-CN-C6H4, 4-NO2-C6H4, 4-Cl-C6H4, 4-Br-C6H4, 4-F-C6H4, 3,4,5-trimethoxy benzyl, 2-pyridyl; R1 = H; R2 is tert-butyl or cyclohexyl and R3 is long aliphatic carbon chain.

3. The compounds with structural formula-(I) as claimed in claim 2 act as antimicrobial agents.