The synthesis of 1-monoacylglycerols of selected unsaturated fatty acids and their antimicrobial and cytotoxicity activity is reported in the present study. The monoacylglycerols of fatty acids like undecenoic, oleic, linoleic and erucic acids were prepared by chemical esterification with solketal followed by deprotection. Fatty acids like alpha linolenic, gamma linolenic and ricinoleic acids were initially isolated from natural sources and further enriched in their respective methyl ester forms. The monoacylglycerols of ricinoleic and linolenic acid methyl esters were prepared by enzymatic transesterification with solketal using lipase from
Structured lipids have gained importance in specialty foods which include modified lipids ranging from glycerides to phospholipids and are produced for specific food, nutritional and biomedical applications. (Osborn and Akoh
Research in this area is gaining importance because of the potential benefits of MAGs such as their biocompatibility and biodegradable nature. There are reports on the higher activity of unsaturated fatty acids compared to saturated fatty acids on some microbial strains (Zheng
Linoleic acid is a nutritionally important fatty acid whose intake is reported to have inverse effects on heart diseases and cholesterol levels (Maryam
Oleic and linoleic acids were purchased from TCI chemicals, India. Solketal, erucic acid and borage oil were purchased from Sigma Aldrich. Eripupae were obtained from the Central Silk Board, Bengaluru, India. Castor oil was purchased from a local market. N,N’-Dicyclohexyl carbodiimide (DCC), 4-Dimethylaminopyridine (DMAP) were obtained from SRL. Penicillin and Streptomycin were used as standard drugs. All solvents were purchased from Merck and used without further purification. The proton NMR spectra were recorded on a Varian 400 MHz instrument and TMS was used as internal standard. Mass spectra were recorded using electron spray ionization on a Waters e2695 Separator module (Waters, Milford, MA, USA) mass spectrometer. IR spectra were recorded in dichloromethane on a Perkin-Elmer Fourier transform (FT)-IR Spectrum BX instrument (Model: Spectrum BX; Connecticut, USA). All the synthesized products were purified by silica gel (60– 120 mesh) column chromatography (Acme Synthetic Chemicals, Mumbai, India) and identified by thin-layer chromatography (TLC). TLC was performed on pre-coated silica gel 60 F254 from Merck (Darmstadt, Germany).
For gamma linolenic acid and alpha linolenic acid, borage oil and eri silkworm oil were chosen as substrates. Both silkworm oil and borage oils were converted to fatty acid methyl esters according to the reported method. Briefly, the oils were refluxed in a 2% sulfuric acid-methanol reagent for 3 hours. After complete conversion as monitored by TLC, the solvent was evaporated and the contents were transferred to a separating funnel and ethyl acetate was added and washed with distilled water until neutral. The organic phase was dried over anhydrous sodium sulphate and concentrated to obtain the fatty acid methyl esters (Christie
Castor oil (15 g) was converted to methyl esters using 300 mL of 2%-sulphuric acid methanol reagent for 3 hours at 75 °C with magnetic stirring. After complete conversion as monitored by TLC, the solvent was evaporated and the contents were transferred to a separating funnel and ethyl acetate (150 mL x2) was added and washed with distilled water until neutral. The organic phase was dried over anhydrous sodium sulphate and concentrated to obtain fatty acid methyl esters. The methyl esters (10 g) were separated into pure methyl ricinoleate (9.04 g; 90% yield) and non-hydroxy fatty acid methyl esters following a reported protocol (Berdeaux
Two temperature programs were employed for the gas chromatographic (GC) analysis. For normal fatty acid methyl esters, GC was performed on an Agilent 6890 gas chromatograph equipped with a flame ionization detector. The column used was a DB-225 with a length of 30 m, 0.25 mm i.d and 25 μm film thickness. The carrier gas was nitrogen at a flow rate of 1 mL/min. The oven programming was as follows: 160 °C for 2 minutes, raised to 230 °C at a rate of 5 °C/min and held at 230 °C for 20 minutes. The injector and detector temperatures were maintained at 220 and 250 °C, respectively. For castor oil fatty acid methyl esters, GC was performed using an Agilent 6890 gas chromatograph coupled with a FID. A HP-1 capillary column (30 m x 0.25 mm x 0.25 μm, (Agilent) was used with a column temperature program of 160°C for 2 min, raised at 10 °C/min to 300 °C and held for 20 min at 300 °C. The injector temperature was 280 °C with a split ratio of 50:1. The carrier gas was nitrogen at a flow rate of 1 mL/min. The detector temperature was 300 °C with air and hydrogen flow rates of 300 mL/min and 30 mL/min, respectively. The fatty acids were identified by comparing the retention times with those of standard fatty acid methyl esters.
The synthesis of 1-monoacylglycerols was carried out in two steps following a reported protocol with slight modifications (Yang
The esterification of oleic acid with solketal is described as an example. In a 100 mL round-bottomed flask, solketal (0.79 g, 0.006 moles) was dissolved in dichloromethane (15 mL) and DMAP (0.2 eq, 0.12 g) was added at 0 ºC. A mixture of DCC (1 eq, 1.03 g) and oleic acid (1 eq, 1.41 g, 0.005 moles) was added and dissolved in 20 mL dichloromethane. The ice bath was removed and the reaction was stirred at room temperature for 24h. After 24 h, the reaction mixture was filtered to remove the precipitate. The filtrate containing the product was washed with saturated aqueous sodium bicarbonate (50 mL) and water (3 x 75 mL) and the organic layer was dried over anhydrous sodium sulphate and concentrated by rotary evaporation at 45 ºC to obtain the crude product. The crude product was purified by silica gel (60–120 mesh) column chromatography using hexane and ethyl acetate gradient to obtain pure acetonide at a 70% yield (1.39 g). The product was concentrated and characterized using 1H-NMR, IR and mass spectral data. A similar protocol was followed for the esterification of undecenoic, linoleic and erucic acids to obtain the corresponding acetonides.
Ricinoleic acid methyl ester (1.5 g; 00 4.8mmol) and solketal (0.69 g; 5.28 mmol) were mixed together. The lipase from
(2, 2-dimethyl-1, 3-dioxolan-4-yl) methyl undec-10-enoate (1a); Yield: 60%; 1H NMR (400 MHz, CDCl3) δ: 1.31 (d, J = 15.7 Hz,-CH3-, 6H), 1.50 – 1.34 (m,-CH2-, 6H), 1.69 – 1.55 (m, -CH2-, 2H), 2.08 – 1.99 (m, CH2-, 2H), 2.34 (t, J = 15.2 Hz,-CH2-, 2H), 3.74 (dd, J = 8.4Hz, 6.2 Hz,-CH-, 1H), 4.13 – 4.05 (m,-CH2- , 2H), 4.17 (dd, J = 11.5Hz, 4.7 Hz, -CH-, 1H), 4.31 (m, -CH-, 1H), 5.06 – 4.89 (m,-CH2-, 2H), 5.81 (m, -CH-, 1H); IR (cm-1): 1741cm-1, 2928, 2855cm-1, 3076cm-1 MS: m/z 321.33 [M+23] .
(2, 2-dimethyl-1, 3-dioxolan-4-yl) methyl oleate (1b); Yield: 70%; 1H NMR (400 MHz, CDCl3) δ: 0.89 (t, J = 6.72Hz, CH3, 3H), 1.34 - 1.26 (m, (CH2)10, 20H), 1.63 - 1.58 (m, CH2, 2H), 2.03 - 1.98 (m, (CH2)2, 4H), 2.36 - 2.32 (t, J = 7.4 Hz, CH2, 2H), 3.75 - 3.71 (m, CH, 1H), 4.18 - 4.05 (m, CH, CH2, 3H), 4.34 – 4.28 (m, CH, 1H), 5.38 - 5.30 (m, (=CH)2, 2H); MS: m/z 419 [M+23]
(9Z,12Z)-(2,2-dimethyl-1,3-dioxolan-4-yl)methyl octadeca-9,12-dienoate (1c); Yield: 90 %; 1H NMR (400 MHz, CDCl3) δ: 0.89 (t, J = 8.2 Hz, CH3, 3H), 1.41 – 1.18 (m,(CH2)9, 18H), 1.48 – 1.40 (m, CH2, 2H), 1.70 – 1.59 (m, CH2, 2H), 2.05 (q, J = 6.8 Hz, CH2, 4H), 2.35 (t, J = 6.95 Hz, 6.4 Hz, CH2, 2H), 2.77 (t, J = 6.5 Hz, 2H), 3.74 (m, CH, 1H), 4.38 – 4.00 (m, (CH2)2, 4H), 5.49 – 5.24 (m ,( CH)4, 4H); IR: 1735cm-1, 2875.50 cm-1, 2966.43 cm-1; MS: m/z = 417.47 [M+23]
(6Z,9Z,12Z)-(2,2-dimethyl-1,3-dioxolan-4-yl)methyl octadeca-6,9,12-trienoate (1d); Yield: 61%; 1H NMR (400 MHz, CDCl3) δ: 0.89 (t, J = 6.91 Hz, CH3, 3H), 1.47 – 1.31 (m, (CH2)6, 12H), 1.74 – 1.59 (m, CH2, 2H), 2.11-2.03 (m, (CH2)2, 4H), 2.36 (t, J = 7.51 Hz, CH2, 2H), 2.96 – 2.63 (m, (CH2)2, 4H), 3.75-3.71 (m, CH, 1H), 4.44 – 3.98 (m, (CH2)2, 4H), 5.49 – 5.25 (m, (CH)6, 6H). IR: 1737 cm-1, 2859 cm-1, 2929 cm-1, 3013 cm-1; Mass: m/z 415.32 [M+23]
(9Z,12Z,15Z)-(2,2-dimethyl-1,3-dioxolan-4-yl)methyl octadeca-9,12,15-trienoate (1e); Yield: 45%; 1H NMR (400 MHz, CDCl3) δ 0.97 (t, J = 7.51 Hz, CH3, 3H), 1.34 – 1.24 (m, (CH2)6, 12H), 1.65 – 1.61 (m, CH2, 2H), 2.09 – 2.03 (m, CH2, 2H), 2.34 (t, J =7.51 Hz, CH2, 2H), 2.86 – 2.73 (m, (CH2)2, 4H), 3.76 – 3.72 (m, CH2, 2H), 4.20 – 4.05 (m, (CH2)2, 4H), 4.37 – 4.29 (m, CH, 1H), 5.44 – 5.27 (m, (=CH)6, 6H); IR: 1265.58 cm-1, 1736.82 cm-1, 2856.53 cm-1, 2930 cm-1, 3054.80 cm-1; Mass: m/z 415.28 [M+23]
(Z)-(2,2-dimethyl-1,3-dioxolan-4-yl)methyl docos-13-enoate (1f); Yield: 82%; 1H NMR (400 MHz) δ: 0.89 (t, J = 6.78 Hz, CH3, 3H) , 1.36 – 1.22 (m, (CH2)16, 32H), 1.66 – 1.60 (m, CH2, 2H), 1.98 - 2.06 (m, CH2, 4H), 2.34 (t, J = 7.58 Hz, CH2, 3H), 3.69 - 3.78 (m, CH, 1H), 4.37 – 3.99 (m, (CH2)2, 4H), 5.39 – 5.31 (m, (=CH)2, 2H); IR: 1733cm-1, 2857 cm-1, 2927 cm-1, 3018 cm-1; MS: m/z 475 [M+23]
(Z)-(2,2-dimethyl-1,3-dioxolan-4-yl)methyl 12-hydroxyoctadec-9-enoate (1g); Yield: 73%; 1H NMR (500 MHz, CDCl3) δ 0.88 (t, J =6.90 Hz, CH3, 3H), 1.58 – 1.53 (m, (CH2)10, 20H), 2.19 – 2.15 (m, (CH2)5, 10H), 2.34 (t, J = 7.58 Hz, CH2, 2H), 3.75 – 3.72 (m, CH, 1H), 4.20 – 4.03 (m, CH2, 2H), 4.35 – 4.27 (m, CH, 1H), 5.57 – 5.37 (m, (CH)2, 2H); IR: 1216 cm-1, 1733cm-1, 2859cm-1, 2930cm-1, 3018cm-1, 3348cm-1; Mass: m/z 435.67 [M+23].
The acetonide of oleic acid (1.25 g, 0.003 moles) dissolved in methanol (40 mL) in a round-bottomed flask was placed in an ice bath. To this cooled mixture, 2 M aqueous hydrochloric acid (2.4 mL, 1.6 eq) were added in one portion. After 15 min, the ice bath was removed and the reaction mixture was stirred for 5 hours at room temperature (20 ºC) while monitoring by TLC at every hour. When the conversion was complete (5 h), the reaction mixture was diluted with 200 mL of diethyl ether, transferred to a separating funnel and washed with 200 mL of saturated aqueous sodium bicarbonate and extracted with diethyl ether (3 x 150 mL). The organic phases containing the product were dried over sodium sulphate and concentrated by rotary evaporation at 45 ºC. The crude product was purified by silica gel column chromatography using a gradient of hexane and ethyl acetate to obtain the corresponding MAG at a 71% yield (0.8 g). The purified product was characterized using 1H-NMR, IR and mass spectral data. The other acetonides in the study were also hydrolyzed following the same methodology to obtain corresponding monoglycerides.
2,3-dihydroxypropyl undec-10-enoate (2a; UDA-MAG): Yield: 73%; H1-NMR (400 MHz, CDCl3) δ: δ 1.49 – 1.18 (m, -CH2, 12H), 2.05 – 2.00 (m, -CH2-, 2H), 2.45 – 2.27 (m, -CH2-, 2H), 3.62 – 3.58 (m, -OH, 1H), 3.71 – 3.68 (m, -OH, 1H), 3.94 – 3.90 (m, CH, 1H), 4.23 – 4.12 (m, -CH2-, 2H), 5.05 – 4.87 (m, -C-H-, 2H), 5.86 – 5.74 (m, -C-H-, 1H); IR: 1729cm-1 , 2928cm-1, 2856cm-1, 3015cm-1, 3407cm-1; MS: m/z 281.38 [M+23]
2, 3-dihydroxypropyl oleate (2b; Oleic-MAG): Yield: 71%; H1-NMR (400 MHz, CDCl3) δ: 0.89 (t, J =6.84 Hz, CH3, 3H), 1.30 – 1.26 (m, (CH2)20, 20H), 1.64 – 1.59(m, CH2, 2H), 2.03 -1.98 (m, (CH2)2, 4H), 2.37 - 2.33 (t, 7.59 Hz, CH2, 2H), 3.71 - 3.57 (m, CH2, 2H), 3.95 -3.82 (m, CH, 1H), 4.22 – 4.12 (m, CH2, 2H), 5.38 – 5.30 (m, (=CH)2, 2H); IR (cm-1): 1730 cm-1, 2851 cm-1, 2926 cm-1, 3010 cm-1, 3404 cm-1; MS: m/z 379.46 [M+23]
(9Z, 12Z)-2, 3-dihydroxypropyl octadeca-9, 12-dienoate (2c; Linoleic-MAG): Yield: 72%; H1-NMR (400 MHz, CDCl3) δ: 0.89 (t, 8.2 Hz, CH3, H), 1.24-1.43 (m, (CH2)7, 14H), 1.59-1.69 (m, CH2, 2H), 2.02-2.11(m, (CH2)2, 4H), 2.35 (t, J = 7.6 Hz, CH2, 2H), 2.77 (t, J = 6.5 Hz, CH, 1H), 3.58-3.72 (m, CH2, 2H), 3.82-3.96(m, CH, 1H) ,4.130 - 4.23 (m, CH2, 2H), 5.30 - 5.42 (m, (CH)4, 4H); IR: 1731.15 cm-1, 2856.27 cm-1, 2927.96 cm-1, 3010.51 cm-1, 3403.57 cm-1; Mass: m/z 377.41 [M+23]
(6Z, 9Z, 12Z)-2, 3-dihydroxypropyl octadeca-6, 9, 12-trienoate (2d; GLA-MAG): Yield: 76%; 1H NMR (400 MHz, CDCl3) δ: 0.89 (t, J = 6.9 Hz, 3H), 1.47 – 1.22(m, (CH2)4, 8H), 1.73 – 1.60 (m, CH2, 2H), 2.11-2.10 (m, (CH2)2, 4H), 2.37 (t, 7.33 Hz, CH2, 2H), 2.89–2.72 (m, (CH2)2, 4H), 3.97-3.56 (m,CH, CH2, 3H), 4.23-4.12 (m, CH2, 2H), 5.49 – 5.23 (m, (CH)6, 6H); IR: 1737 cm-1, 2857 cm-1, 2927 cm-1, 3011 cm-1, 3382 cm-1; Mass: m/z 375.45 [M+23].
(9Z, 12Z, 15Z)-2,3-dihydroxypropyl octadeca-9,12,15-trienoate (2e; ALA-MAG): Yield: 72%; 1H NMR (400 MHz, CDCl3) δ : 0.97 (t, 7.47Hz, CH3, 3H), 1.41 – 1.27 (m, (CH2)4, 8H), 1.64 – 1.60 (m, CH2, 2H), 2.11 – 2.03 (m, (CH2)2, 4H), 2.35 (t, 7.47 Hz, CH2, 2H), 2.90 – 2.72 (m, (CH2)2, 4H), 3.78 – 3.50 (m, CH2, 2H), 4.00 – 3.86 (m, CH, 1H), 4.22 – 4.13 (m, CH2, 2H), 5.50 – 5.18 (m, (CH)6, 6H); IR: 1265.18 cm-1, 1729 cm-1, 2928.70 cm-1, 3056.94 cm-1, 3383.57 cm-1; Mass: m/z 375.25 [M+23]
(Z)-2,3-dihydroxypropyl docos-13-enoate (2f; Erucic-MAG): Yield: 90%; 1H NMR (500 MHz) δ 0.88 (t, J = 6.9 Hz, CH3, 3H), 1.38 – 1.20 (m, (CH2)14, 28H), 1.60-1.66 (m, CH2, 2H), 1.99 - 2.01 (m, (CH2)2, 4H), 2.35 (t, J = 7.6 Hz, CH2, 2H), 3.72 – 3.58 (m, CH2, 2H), 3.96 – 3.90 (m, CH, 1H), 4.18 (m, CH2, 2H), A5.50 – 5.09 (m, (=CH)2, 2H); IR: 1730 cm-1, 2856 cm-1, 2927 cm-1, 3021 cm-1, 3418 cm-1; Mass: m/z = 435 [M+23]
(Z)-2,3-dihydroxypropyl 12-hydroxyoctadec-9-enoate (2g; Ricinoleic-MAG): Yield: 80%; 1H NMR (400 MHz, CDCl3) δ 0.89 (t, J = 6.8 Hz, CH3, 3H), 1.41 – 1.22 (m, (CH2)8, 16H), 1.47 (t, J = 6.0 Hz, CH2, 2H), 1.69 – 1.57 (m, CH2, 2H), 2.08 – 2.03 (m, CH2, 2H), 2.21 (t, J = 6.3 Hz, CH2, 2H), 2.35 (t, 7.45 Hz, CH2, 2H), 3.66 – 3.56 (m, CH2, 2H), 3.91 – 3.98 ( m, CH, 1H), 3.96 – 3.91 (m, CH, 1H), 4.23 – 4.13 (m, 2H, 2H), 5.63 – 5.32 (m, (=CH)2, 2H); IR: 1216 cm-1, 1728 cm-1, 2858 cm-1, 2929 cm-1, 3017 cm-1, 3389 cm-1; Mass: m/z 395.22 [M+23].
The in vitro antibacterial activity of the newly synthesized compounds was studied against the bacterial strains,
The minimum inhibitory concentrations (MIC) of various synthetic compounds were tested against three representative Gram-positive organisms viz.
The cell viability (MTT test) of the different test compounds was assessed following our earlier published literature (Kalpana
Statistical analysis was carried out using Graph Pad Prism version 6.04 for Windows, Graph Pad Software, San Diego. The data were analyzed with unpaired Student’s t-test, followed by ‘‘One-way ANOVA’’ by the Dunnett’s multiple comparisons test.
This study was intended to evaluate the antimicrobial and cytotoxic behavior of the 1-monoacylglycerols of selected unsaturated fatty acids. Among the selected fatty acids, GLA and ALA were obtained from urea complexation from borage and eri silkworm oils, respectively, whereas other fatty acids were commercially available. Eri silkworm oil and borage oil are reported to contain high amounts of ALA and GLA, respectively, and hence were taken as source oils for PUFAs. Urea complexation was carried out repeatedly to obtain maximum enrichment of the unsaturated fatty acids in the study. Ricinoleic acid was obtained from castor oil in pure form by selective extraction of hydroxy fatty acid using hexane and aqueous methanol solvent mixtures.
Fatty acid composition (wt%) of substrate oils and their enriched fractions
Fatty acid | SWO | ALA-rich fraction from SWO | Borage oil | GLA-rich fraction from Borage oil | Castor oil | Ricinoleic acid-rich fraction from castor oil |
---|---|---|---|---|---|---|
14:0 | 0.3 ± 0.06 | - | - | - | - | - |
16:0 | 24.9 ± 0.94 | - | 10.0 ±0.03 | - | 1.3 ±0.33 | - |
16:1 | 1.3 ± 0.16 | - | 0.1 ±0.01 | - | - | - |
18:0 | 3.8 ± 0.31 | - | 4.2 ±0.03 | - | 1.2 ±0.32 | - |
18:1 | 13.2 ± 0.27 | 1.1 ±0.01 | 17.0 ±0.07 | - | 4.3 ±1.01 | 0.3 ±0.03 |
18:2 | 4.2 ± 0.04 | 4.7 ±0.13 | 37.3 ±0.09 | 4.1 ±0.09 | 5.0 ±0.42 | 0.4 ±0.04 |
18:1-O | - | - | - | - | 88.2 ±2.08 | 99.3 ±0.07 |
18:3-A | 51.6 ±0.57 | 94.1 ±0.13 | 0.3 ±0.04 | 1.3 ±0.06 | - | - |
18:3-G | - | - | 22.4 ±0.03 | 94.6 ±0.04 | - | - |
20:0 | - | - | 0.2 ±0.01 | - | - | - |
20:1 | - | - | 4.1 ±0.06 | - | - | - |
22:0 | - | - | 0.2 ±0.01 | - | - | - |
22:1 | - | - | 2.6 ±0.05 | - | - | - |
24:0 | - | - | 1.5 ±0.01 | - | - | - |
It can be observed that in both silkworm and borage oils, the PUFA fraction was selectively enriched with urea complexation, which is usually the method of choice for PUFA concentration (Hayes
The synthesis of 1-monoacylglycerols of unsaturated fatty acids in the present study was carried out in two steps. Initially the fatty acids were esterified to solketal employing DCC as the coupling reagent and DMAP as the catalyst, which is a wellknown methodology for esterification (Yang
Synthetic route for 1-monoolein.
As fatty acids like ALA and GLA were enriched in their methyl ester form, an enzymatic transesterification route was found to be more suitable to avoid the hydrolysis of methyl esters and subsequent chemical esterification steps. This has not only reduced the number of steps but also provided an opportunity to include an enzymatic step which was a simple, mild and green methodology. The lipase employed for this transesterification was an immobilized lipase from
Synthetic route for 1-monoricinolein.
The procedures using DCC/DMAP for esterification and the hydrolysis of acetonide with HCl were mild and the yields were also satisfactory. The products of esterification and deprotection were purified by column chromatography and characterized by IR, NMR and mass spectral data. The structures of the prepared 1-MAGs are shown in
Structures of the 1-monoacylglycerols.
The synthesized 1-MAGs were subjected to antimicrobial evaluation against both bacterial and fungal strains and also for cytotoxicity assay against five cell lines. It was found that the MAGs showed antibacterial action but were not found to exhibit antifungal activity. The antibacterial activity was tested against both Gram-positive and Gram-negative bacterial strains. The results of the antibacterial activity were measured as zone of inhibition (in mm) and are presented in
Bacterial zone of inhibition (mm) of test compounds
Name of the Compound with code | Staphylococcus aureus | Staphylococcus epidermidis | Bacillus subtilis | Escherichia coli | Pseudomonas aeruginosa | Klebsiella pneumoniae |
---|---|---|---|---|---|---|
1-Monolaurin (lipid reference standard) | 15.8±0.7 | - | 18.8±0.7 | 15±1 | 18.33±0.5 | - |
11±0.5 |
- | 8±1 |
- | 19.66±0.5 |
- | |
- | - | - | - | - | - | |
- | - | - | - | - | - | |
18.6±0.5 |
19.16±0.2 |
22.5±0.5 |
18.6±0.5 |
20.33±1.1 |
- | |
16.8±0.2 |
- | 20.8±0.2 |
- | - | - | |
- | 20±0.5 |
- | - | - | - | |
11.6±0.5 |
- | - | - | - | - | |
Penicillin (standard) | 28±0.2 |
25.83±0.28 | 27.6±0.5 |
24.16±0.7 |
23.16±0.28 |
25.33±0.5 |
Streptomycin | 22.8±0.2 |
25.16±0.2 | 24±1 |
23.3±0.5 |
27.33±0.5 |
26.16±0.2 |
±, Mean value with standard error of mean of triplicates; ‘‘One-way ANOVA’’ used to measure the mean differences among the compounds by the Dunnett’s multiple comparisons test.
Significantly different from (lipid reference standard, 1-Monolaurin) at P < 0.001;
Significantly different from Penicillin at P < 0.001;
Significantly different from (lipid reference standard, 1-Monolaurin) at P < 0.05;
notsignificant
It was observed that most of the prepared 1-MAGs were more effective against Gram-positive bacteria compared to Gram-negative bacteria. Among the lipids, GLA-MAG showed a broad spectrum antibacterial activity against all the tested strains except against
Minimum Inhibitory Concentration (μg/mL) of test compounds
Compound | Staphylococcus aureus | Staphylococcus epidermidis | Bacillus subtilis | Escherichia coli | Pseudomonas aeruginosa | Klebsiella pneumoniae |
---|---|---|---|---|---|---|
150 | >150 | 150 | >150 | 9.37 | >150 | |
>150 | >150 | >150 | >150 | >150 | >150 | |
>150 | >150 | >150 | >150 | >150 | >150 | |
18.75 | 37.5 | 4.74 | 37.5 | 9.37 | >150 | |
37.5 | >150 | 9.37 | >150 | >150 | >150 | |
>150 | 9.37 | >150 | >150 | >150 | >150 | |
150 | >150 | >150 | >150 | >150 | >150 | |
1-Monolaurin | 37.5 | >150 | 18.75 | 37.5 | 18.75 | >150 |
Penicillin | 1.562 | 3.125 | 1.562 | 12.5 | 12.5 | 6.25 |
Streptomycin | 6.25 | 3.125 | 6.25 | 6.25 | 1.562 | 3.125 |
Media values are represented in μg/mL of serial dilution of two replicates. For all the test compounds, the highest concentration was used at 150 μg/mL and lowest concentration used at 2.34 μg/mL; whereas, for standards (Penicillin and Strepotomycin) the highest concentration used at 100 μg/mL and lowest concentration at 0.78 μg/mL. Values with > 150 are considered to show no antibacterial activity.
After the MAG of GLA, the next active MAG was observed to be of ALA, which showed inhibition against
The cytotoxicity evaluation for the prepared MAGs was carried out by the widely employed MTT assay and the results are presented in
in vitro cytotoxicity test of compounds (μM) against cancerous and non-cancerous cell lines
S.No | Test compound | IC50 values (μM) |
||||
---|---|---|---|---|---|---|
HeLa | B16-F10 | SKOV3 | MCF-7 | CHO-K1 | ||
1 | 35.62 ± 0.20 | 42.12 ± 0.08 | 31.10 ± 0.18 | 15.32 ± 0.05 | 73.29 ± 2.2 | |
2 | 28.35 ± 0.28 | 97.36 ± 0.10 | 21.89 ± 0.08 | 23.03 ±0.01 | 30.03 ± 1.7 | |
3 | 32.13 ± 0.15 | 45.25 ± 0.96 | 27.34 ± 0.85 | 16.60 ± 0.11 | 43.32 ±0.28 | |
4 | 27.54 ± 0.38 | 44.04 ± 0.15 | 53.53 ± 2.2 | 16.42 ± 0.21 | 39.93 ± 0.23 | |
5 | 25.55 ± 0.58 | 62.93 ± 0.83 | 26.75 ± 0.10 | 13.32 ± 0.15 | 28.97 ± 0.27 | |
6 | 23.98 ± 0.79 | 52.96 ± 0.72 | 44.65 ± 0.60 | 13.91 ± 0.01 | 51.61 ± 0.12 | |
7 | 24.61 ± 0.37 | 99.20 ± 2.0 | 20.55 ± 0.77 | 15.60 ± 0.07 | 33.02 ± 0.44 | |
8 | 1-Monolaurin | 25.60 ± 0.15 | 69.79 ± 1.3 | 31.35 ± 0.03 | 15.64 ± 0.06 | 50.25 ± 1.3 |
|
0.8 ± 0.71 | 0.7 ± 0.56 | 0.8 ± 0.63 | 2.0 ± 0.81 | NA | |
Mitomycin C | NA | NA | NA | NA | 13.1 ± 0.68 |
Cancer cell lines: (HeLa, Homo sapiens cervix adenocarcinoma; B16-F10, Mouse skin melanoma (ATCC® CRL 6475™); SKOV3, Human Ovarian cancer (ATCC® HTB 77™); MCF7, Human Breast Adenocarcinoma (ATCC® HTB-22™); Non-cancerous cell lines (CHO-K1, Chinese Hamster Ovary cells (ATCC® CCL-61™); ±, Standard deviation of mean value of triplicates using Graph Pad Prism software; NA, no activity.
The present study was helpful in assessing the antimicrobial and cytotoxicity potential of the 1-MAGs of some of the unsaturated fatty acids and further studies on a range of 1-MAGs of different fatty acids could be of interest in view of developing biocompatible biologically active agents.
The synthesis of structured 1-monoacylglycerols comprising unsaturated fatty acids and their antimicrobial and cytotoxicity efficacies were studied. Among the prepared lipid derivatives, the monoacylglycerol of gamma linolenic acid was found to be the most effective against both Gram-positive and Gram-negative bacterial strains. All the MAGs prepared exhibited moderate cytotoxicity activity against a panel of cell lines. However, further studies are needed to exploit the full potential of these monoacylglycerol derivatives with different types of fatty acids to test in food-based systems for additional applications.
Ms Juliya Johny and Dr. Shiva Shanker Kaki gratefully acknowledge the Science and Engineering Research Board, Department of Science & Technology, New Delhi, for financial assistance under the Early Career Research Award [project file number ECR12017/000639] and the Director, CSIR-IICT, for providing the facilities.
IICT Communication number: IICT/Pubs./2018/291.