Conoidecyclics A-C from marine macroalga Turbinaria conoides: Newly described natural macrolides with prospective bioactive properties Author Chakraborty, Kajal * & Marine Bioprospecting Section of Marine Biotechnology Division, Central Marine Fisheries Research Institute, Ernakulam North, P. B. No. 1603, Cochin, Kerala State, & * Author Dhara, Shubhajit text Phytochemistry 2021 112909 2021-11-30 191 1 14 http://dx.doi.org/10.1016/j.phytochem.2021.112909 journal article 264237 10.1016/j.phytochem.2021.112909 8fb5ad8b-262f-4221-b858-8817e02af56a 1873-3700 8258118 4.3. Chromatographic purification of organic extract of T. conoides The solvent extract of T. conoides ( 30 g ) was fractionated by sequential chromatographic procedures. The crude was mixed with the silica gel (60–120 mesh, 3.0 g) and loaded into a column of glass ( 0.1 m × 40 mm ) with silica gel (60–120 mesh, 50 g ). The elution was initiated with less polar solvent ( n -hexane), followed by an increase of polarity (EtOAc to MeOH) to collect twenty-five fractions (10 mL/fraction), which were combined to five (TC 1 through TC 5 ) ( Table S1 ) based upon TLC (8:2 v/v, n -hexane:EtOAc) and RP C 18 -HPLC (MeOH/acetonitrile MeCN, 3:2 v/v) analysis, and those fractions (TC 1 -TC 5 ) were assessed for their bioactivities. The fractions possessing potential bioactive properties were chosen for further purification. The fractions TC 3 (EtOAc: n - hexane 2:3 v/v; 4.65 g , 15.5% yield) and TC 4 (EtOAc: n -hexane 7:3 v/v; 4.26 g , 14.2% yield) displayed greater attenuation potential against COX-2 and 5-LOX enzymes. The radical scavenging potential of TC 3 and TC 4 against 1,1-diphenyl-2-picryl-hydrazil (DPPH) and 2, 2 ′ -azino-bis-3- ethylbenzothiozoline-6-sulfonic acid (ABTS + ) were found to be greater compared to others ( Table S1 ), and therefore, were selected for downstream purification. Likewise, the sub-fraction TC 3 was subjected to fractionation on a silica gel (230–400 mesh) loaded glass column ( 450 mm × 30 mm ). Step-wise gradient elution with n -hexane-EtOAc (9:1 to 3:2, v/v) yielded twenty sub-fractions (20 mL/fraction), which were combined to four (TC 3-1 -TC 3-4 ) after TLC ( n -hexane-EtOAc, 4:1 v/v) and RP-C 18 HPLC (MeOH–MeCN, 3:2 v/v) experiments. The fraction TC 3-4 ( 1.19 g , 3.96% yield) eluted at n -hexane-EtOAc (6:4, v/v) displayed potential bioactivities ( Table S1 ). Therefore, TC 3-4 was further subfractionated on fine silica gel (SP 1 –B 1A , 230–400 mesh, 12 g ) loaded into a column connected to the flash chromatography system (SP 1 –B 1A , Biotage, Sweden ) by increasing gradient of n -hexane/EtOAc/MeOH to acquire twelve sub-fractions (15 mL/fraction). The latter fractions were combined to three (TC 3-4-1 -TC 3-4-3 ) after TLC ( n -hexane-EtOAc, 8:2 v/v) and RP-C 18 HPLC (MeOH/MeCN, 3:2 v/v). TC 3-4-1 ( 0.768 g , 2.56% yield) was purified on silica gel GF 254 by n -hexane: EtOAc (25:1, v/v) solvent system using the preparatory TLC (PTLC) system to yield conoidecyclic A ( 85 mg , 0.28%, based on dry weight) and conoidecyclic B ( 73 mg , 0.24%, based on dry weight), and their homogeneity was confirmed by TLC (EtOAc/ n -hexane, 1:9 v/v) and RP-C 18 HPLC (MeOH/ MeCN, 3:2 v/v). The fraction TC 4 was subjected to flash chromatography on a silica gel column (230–400 mesh) with a gradient elution of n -hexane/EtOAc/MeOH to yield sixteen fractions (15 mL/fraction), which were combined into three (TC 4-1 -TC 4-3 ) after TLC (EtOAc- n -hexane, 1:4 v/v). TC 4-2 ( 1.15 g , 3.83% yield) displayed greater bioactive potential than other column sub-fractions ( Table S1 ), and flash chromatographic separation using a gradient elution of n -hexane/EtOAc yielded eleven fractions (13 mL/fraction), which were combined into three (TC 4-2-1 -TC 4-2-3 ). The bioactive fraction TC 4-2-3 ( 0.442 g , 1.47% yield) was a mixture, and was further purified by PTLC (EtOAc: n -hexane, 1:26, v/v) to obtain conoidecyclic C ( 65 mg , 0.22%, based on dry weight). The homogeneity of the latter was confirmed by RP-C 18 HPLC (MeOH–MeCN 3:2, v/v) and TLC (EtOAc/ n -hexane 1:4, v/v).