The as-prepared carbodithioate derivative 3 is considered to be a key molecule for the preparation of new antipyrine-1,3,4-thiadiazole-based molecules (4–9) through its reaction with the appropriate hydrazonoyl halides. Furthermore, a typical Biginelli three-component cyclocondensation reaction involving ethyl acetoacetate, 4-antipyrinecarboxaldhyde, and thiourea under the standard conditions is carried out in the presence of sulfuric acid to afford the corresponding antipyrine–pyrimidine hybrid molecule (10). The latter was submitted to react with hydrazine monohydrate to provide the corresponding hydrazide derivative (11) which, under reaction with ethyl acetoacetate in refluxing ethanol containing catalytic amount of acetic acid, afforded the corresponding derivative (12). The structure of the newly synthesized compounds was affirmed by their spectral and microanalytical data. We also screened for their antimicrobial potential (ZOI and MIC) and conducted a kinetic study. Additionally, the mechanism of biological action was assessed by a membrane leakage assay and SEM imaging technique. Moreover, the biological activities and the binding modes of these compounds were further supplemented by an in silico docking study against E. coli β-carbonic anhydrase. The amount of cellular protein released by E. coli is directly correlated to the concentration of compound 9, which was found to be 177.99 µg/mL following treatment with 1.0 mg/mL of compound 9. This finding supports compound 9’s antibacterial properties and explains how the formation of holes in the E. coli cell membrane results in the release of proteins from the cytoplasm. The newly synthesized compounds represent acceptable antimicrobial activities with potential action against E. coli β-carbonic anhydrase. The docking studies and antimicrobial activity test proved that compound (9) declared a greater activity than the other synthesized compounds.In this investigation, 4-antipyrinecarboxaldhyde was reacted with methyl hydrazinecarbodithioate to afford the carbodithioate derivative  Antipyrine-based molecules continue to be of great interest for both medicinal chemists and organic chemists owing to their richness in synthetic compounds with multifarious involvement in pharmaceutical and medicinal compounds ,2. In thOn the other hand, 1,3,4-thiadiazoles ,15,16,17E. coli, particularly among immunocompromised individuals utilizing urethral and intravascular catheters  at m/z = 498 in an approximate ratio of 1:1, which supported the molecular formula (C23H21ClN6O3S).4-antipyrinecarboxaldhyde (oate (3) . We wereperature . The che1), and thiourea to afford the desired molecule 10. In the reaction performed via the tetrahedral mechanism, in which the N-C bond was formed first and then the C-O bond begin to break, the elimination of ethanol occurred, and so a lot of energy was accumulated in the medium of the reaction to offset the activation energy of the reaction, and a conversion took place, followed by cyclization via the nucleophilic attack through the amine group onto the carbonyl group to give the target molecule 10. Compound 10 was reacted with hydrazine hydrate to yield the corresponding hydrazide derivative 11, which then reacted with ethyl acetoacetate in ethanol containing a catalytic amount of acetic acid to afford the corresponding pyrazole derivative 12 [12 was confirmed based on correct spectral data, in which its 1H-NMR spectrum revealed three singlet signals at δ 1.94, 2.26, 3.34 ppm for three methyl groups, a doublet signal at δ 3.12 ppm for protons of (CH2-pyrazole), a singlet signal at δ 3.95 ppm for the proton of , a multiplet signal in the region from δ 7.32 to 7.52 ppm for the aromatic protons, and two singlet signals at δ 8.25 and 10.01 ppm for the NH protons. Its 13C-NMR spectrum showed significant signals at δ 15.72 ppm for (CH3), 17.94 ppm for (CH3), 42.15 ppm for (CH2-pyrazole), 51.22 ppm for , 106.14, 108.24, 120.12, 121.61, 122.12, 123.01, 134.57, 143.09, 152.98, and 159.14 ppm, which represented the aromatic carbons, 163.24 ppm for (C=O), 164.58 ppm for (C=O), 165.19 ppm for (C=O), and 182.12 ppm for (C=S).Additionally, a new polysubstituted pyrimidine derivative containing antipyrine moiety was constructed via the Biginelli three-component cyclocondensation reaction ,44,45,46ative 12   had the greatest effect on E. coli. With inhibition zones of 16.9 mm and 19.0 mm, it had the strongest effect on C. albicans of all the studied fungal strains. Additionally, compound 9 showed potential antibacterial action against E. coli, P. aeruginosa, S. aureus, B. subtilis, C. albicans, and C. neoformans, with inhibition zones of 19.0, 14.2, 17.0, 15.0, and 16.9 mm, respectively, at a concentration of 100 µg/mL (100 ppm). The largest effect was against E. coli and S. aureus with inhibition zones of 14.0 and 13.1 mm, respectively, for compound 4 and 17.5 and 11.3 mm for compound 5 against E. coli and B. subtilis. However, compounds 4 and 5 possessed antibacterial activity that was less potent than compound 9. Additionally, the MICs of every sample that was evaluated  were established, as shown in The results showed that compound 9, 5, and 4 had the best MICs towards the tested bacteria and unicellular fungus, with MICs in the range of 15.62–500 µg/mL. Additionally, E. coli was shown to be the most susceptible of the studied bacteria, with the MICs of compounds 9, 5, and 4 being 15.62, 31.25, and 125 µg/mL, respectively. On the other hand, the MIC of every substance against E. coli, P. aeruginosa, and S. aureus ranged from 31.25 to 500 µg/mL, which was lower than that of B. subtilis.The results showed that, when compared to other examined microbiological strains, compounds E. coli) and unicellular fungi . On the other hand, the utilized standard antibiotics (AMC/Nyst) were more active than the synthesized compound against some tested microbes due to the fact that the concentration used in our synthesized compound 9 , the proposed compounds showed good antibacterial efficacy against some bacterial isolates , compound .D. data , compounE. coli bacteria. According to E. coli is directly correlated to the concentration of compound 9, which was found to be 177.99 µg/mL following the treatment of compound 9 with 1.0 mg/mL. This finding supports compound 9’s antibacterial properties and explains how the formation of holes in the E. coli cell membrane results in the release of proteins from the cytoplasm.The Bradford technique was used to calculate the amounts of protein released in the suspension of the treated 9 made E. coli cell membranes more permeable; therefore, it was plausible to anticipate that membrane permeability would be mistaken for an essential factor in the reduction in bacterial mass. According to these test results, compound Similar results were found when ferrites were used, as described in related studies, which showed concentration-dependent membrane instability in bacterial cells and indicated intracellular substance leakage .Paul et al.  demonstrE. coli. As shown in 9 revealed bacterial groups that generally prolonged and grew with a regular surface, a normal shape, and a normal count.9 (the most active compound), E. coli displayed striking morphological abnormalities  nano-drug, on the other hand, was subjected to SEM imaging against 2 hydration. In addition, they play a pivotal role in many cellular and physiological processes. In the present study, an in silico docking technique is used to investigate the binding mode of actions of the newly synthesized compounds and a reference drug against the target enzyme E. coli β-carbonic anhydrase. From the data obtained, the newly compounds displayed respectable fitting to the active site residues of the enzyme, which showed binding energies ranging from −9.7 to −7.0 kcal/mol. Compound 9 unveiled the best binding energy  and showed H-bond and arene–sigma interactions with the residues TRP64 and TYR63, respectively. On the other hand, compound 3 showed the lowest binding energy  against the target enzyme, as represented in Carbonic anhydrases CAs are important metalloenzymes widely present in all living organisms and catalyze the essential reaction of CO9, as an example, were calculated using the ADMET lab tool, by plotting radar showing 13 properties 2SO solutions on a BRUKER 500 FT-NMR system spectrometer , and chemical shifts were expressed in ppm units using TMS as an internal reference. Mass spectra were recorded on a GC-MS QP1000 EX Shimadzu . Elemental analyses were carried out at the Microanalytical Center of Cairo University, Egypt.All melting points were uncorrected and measured using an electrothermal device. The IR spectra were recorded (KBr discs) using the Shimadzu FT-IR 8201 PC spectrophotometer . 1  and the methyl hydrazinecarbodithioate 2  in 20 mL isopropyl alcohol was stirred at room temperature for 2 h, and the reaction was monitored by TLC. The solid was collected and washed with (water/ethanol) the recrystallized from the ethanol to give the desired derivative. Yellow crystals (85%); m.p.: 200–202 °C, FT-IR : v 3296 (N-H), 1664 (N-C=O), 1620 (C=N), 1595 (C=C); 1H-NMR (DMSO-d6): δ 2.47 , 2.60 , 3.26 , 7.30–7.51 , 8.05 , 13.03 ;13C-NMR : δ 12.22 (CH3), 18.48 (S-CH3), 34.09 (N-CH3), 123.70 (Ar), 123.88 (Ar), 125.82 (Ar), 127.82 (Ar), 129.26 (Ar), 132.82(Ar), 151.53(Ar), 155.18(CH), 164.41(C=O), 195.81 (C=S); MS m/z (%): 320 . Anal. Calcd. for “C14H16N4OS2” (320): C, 52.48; H, 5.03; N, 17.49 Found: C, 52.42; H, 5.01; N, 17.43%.A mixture of 4-antipyrinecarboxaldhyde 3  and the selected derivatives of the hydrazonoyl halides (5 mmol) and 2–3 drops of diisopropylethylamine (DIPEA) as a catalyst in 10 mL ethanol was stirred at room temperature for 2 h. The stirring continued for approximately 5 h, and the reaction was monitored by TLC. The solid was collected and washed with (water/ethanol) the recrystallized from the proper solvent to give the desired derivatives 4–9, respectively.A mixture of compound 4--ylidene)hydrazono)methyl)-1,5-dimethyl-2-phenyl-1,2-dihydro-3H-pyrazol-3-one (4), yellow crystals from acetic acid (78%); m.p. 260–261 °C, FT-IR : 1684 (C=O), 1675 (N-C=O), 1610 (C=N), 1580 (C=C); 1H-NMR (DMSO-d6): δ 2.42 , 2.56 , 3.22 , 7.27–7.7.96 , 8.09 ; 13C-NMR : δ 12.34 (CH3), 24.28 (CH3), 34.21 (N-CH3), 94.12 (Ar), 121.32 (Ar), 121.72 (Ar), 122.50 (Ar), 123.70 (Ar), 123.88 (Ar), 125.82 (Ar), 127.82 (Ar), 129.26 (Ar), 132.82(Ar), 133.12(Ar), 134.22 (Ar), 134.82 (Ar), 151.53 (Ar), 158.18 (C=N), 163.7 (H-C=N), 164.41 (C=O), 195.81 (C=O); MS m/z (%): 432 . Anal. Calcd. for “C22H20N6O2S” (432): C, 61.10; H, 4.66; N, 19.43; Found: C, 61.12; H, 4.61; N, 19.38%.4-(5-acetyl-3-(4-nitrophenyl)-1,3,4-thiadiazol-2(3H)-ylidene)hydrazono)methyl)-1,5-dimethyl-2-phenyl-1,2-dihydro-3H-pyrazol-3-one (5), yellowish brown crystals from acetic acid (62%); m.p. 245–247 °C, FT-IR : 1686 (C=O), 1672 (N-C = O), 1600 (C=N), 1580 (C=C); 1H-NMR (DMSO-d6): δ 2.36 , 2.48 , 3.22 , 7.06–7.47 , 7.94 ; 13C-NMR : δ 12.36 (CH3), 24.18(CH3), 34.22(N-CH3), 95.22 (Ar), 120.82 (Ar), 122.72 (Ar), 122.98 (Ar), 123.78 (Ar), 123.18(Ar), 124.62 (Ar), 128.52 (Ar), 129.16 (Ar), 132.52 (Ar), 133.22 (Ar), 134.12 (Ar), 134.82 (Ar), 151.23 (Ar), 158.18 (C=N), 163.18 (H-C=N), 163.41 (C=O), 197.81(C=O); MS m/z (%): 477 . Anal. Calcd. for “C22H19N7O4S” (477): C, 55.34; H, 4.01; N, 20.53; Found: C, 55.29; H, 3.97; N, 20.46%.Ethyl 4-(4-chlorophenyl)-5-methylene)hydrazono)-4,5-dihydro-1,3,4-thiadiazole-2-carboxylate (6), yellow crystals from acetic acid (82%); m.p. 175–177 °C, FT-IR : 1675 (N-C = O), 1715(C=O), 1615 (C=N), 1600 (C=C); 1H-NMR (DMSO-d6): δ 1.31, 2.63 , 3.30 , 4.36, 7.34–7.60 , 8.19 ; 13C-NMR : δ 12.33 (CH3), 12.35 (CH23CH), 13.93 (N-3CH), 62.77 (2CHCH3), 86.40 (Ar), 99.75 (Ar), 123.25 (Ar), 125.78 (Ar), 127.67(Ar), 129.00(Ar), 129.19 (Ar), 133.19 (Ar), 134.48 (Ar), 138.20 (Ar), 149.95 (Ar), 152.86 (Ar), 158.30(Ar), 163.30 (CH), 163.71 (C=O), 168.72 (C=O); MS m/z (%): 498 , 496 . Anal. Calcd. for “C23H21ClN6O3S” (496): C, 55.59; H, 4.26; N, 16.91; Found: C, 55.59; H, 4.26; N, 16.91%.Ethyl-5-methylene)hydrazono)-4-(4-nitrophenyl)-4,5-dihydro-1,3,4-thiadiazole-2-carboxylate (7), yellowish brown crystals from dioxane (76%); m.p. 157–159 °C, FT-IR : 1668 (N-C=O), 1725 (C=O), 1600 (C=N), 1560 (C=C); 1H-NMR (DMSO-d6): δ 1.30 , 2.48 , 3.29 , 4.33 , 7.33–7.57 , 8.35 ; 13C-NMR : δ 12.28 (CH3), 12.37 (CH23CH), 13.95 (N-3CH), 61.97 (2CHCH3), 86.40 (Ar), 123.27 (Ar), 125.18(Ar), 127.77(Ar), 129.12(Ar), 129.16 (Ar), 133.18 (Ar), 134.28 (Ar), 138.22 (Ar), 149.15 (Ar), 151.96 (Ar), 157.33 (Ar), 163.32 (CH), 163.77 (C=O), 165.82 (C=O); MS m/z (%): 507. Anal. Calcd. for “C23H21N7O5S” (507): C, 54.43; H, 4.17; N, 19.32; Found: C, 54.47; H, 4.12; N, 19.28%.5-methylene)hydrazono)-N-phenyl-4-(p-tolyl)-4,5-dihydro-1,3,4-thiadiazole-2-carboxamide (8), yellow crystals from ethanol (68%); m.p. 260–262 °C, FT-IR : 3315(NH), 1662 (N-C = O), 1680 (C=O), 1615 (C=N), 1598 (C=C); 1H-NMR (DMSO-d6): δ 2.31, 2.45 , 3.32 , 7.31–8.06 , 8.15 , 10.63 ; 13C-NMR : δ 12.36 (CH3), 20.51 (CH3), 34.06 (N-CH3), 100.12 (Ar), 120.89 (Ar), 121.72 (Ar), 124.50 (Ar), 125.62(Ar), 127.52(Ar), 128.58 (Ar), 129.12(Ar), 129.20(Ar), 146.61(Ar), 147.00 (Ar), 148.63 (Ar), 156.24 (C=O), 162.24 (CH), 163.45(C=O); MS m/z (%): 507. Anal. Calcd. for “C28H25N7O2S” (523): C, 64.23; H, 4.81; N, 18.73; Found: C, 64.19; H, 4.78; N, 18.68 %.5-methylene)hydrazono)-4-(4-nitrophenyl)-N-phenyl-4,5-dihydro-1,3,4-thiadiazole-2-carboxamide (9), brown crystals from ethanol (63%); m.p. 281–283 °C, FT-IR : 3322(NH), 1668 (N-C=O), 1680(C=O), 1615 (C=N), 1600 (C=C); 1H-NMR (DMSO-d6): δ 2.50 , 3.28 , 7.21–7.78 , 8.08 , 10.79 ; 13C-NMR : δ 12.36 (CH3), 32.12 (N-CH3), 98.14 (Ar), 121.89 (Ar), 122.74 (Ar), 123.51 (Ar), 124.52 (Ar), 126.82 (Ar), 127.61(Ar), 128.15 (Ar), 128.28 (Ar), 145.62 (Ar), 146.50 (Ar), 149.83 (Ar), 155.24(C=O), 163.24 (CH), 163.55 (C=O); MS m/z (%): 554 . Anal. Calcd. for “C27H22N8O4S” (554): C, 58.48; H, 4.00; N, 20.21; Found: C, 58.42; H, 3.95; N, 20.17%.Ethyl 4--6-methyl-2-thioxo-1,2,3,4-tetrahydropyrimidine-5-carboxylate (10): A mixture of 1,5-dimethyl-3-oxo-2-phenyl-2,3-dihydro-1H-pyrazole-4-carbaldehyde (1) , ethyl acetoacetate , and thiourea  was dissolved in 15 mL DMF with the addition of a few drops of sulfuric acid. The reaction mixture was heated under reflux for 5hrs. After cooling, the solid formed was separated and recrystallized from dioxane as yellow crystals; Yield (83%); m.p. 212–214 °C, FT-IR : 3415, 33821(NH), 668 (N-C=O), 1735(C=O), 1620 (C=N), 1600 (C=C); 1H-NMR (DMSO-d6): δ 1.02, 2.17, 3.23 , 3.50, 3.96, 7.29–7.45 , 12.23 , 12.29 ; 13C-NMR : δ 13.57 (CH23CH), 15.58 (CH3), 18.54 (CH3), 52.02 , 61.66 (2CHCH3), 103.84 (Ar), 106.44 (Ar), 120.20(Ar), 124.41(Ar), 126.92(Ar), 134.47(Ar), 153.09 (Ar), 160.98 (Ar), 166.18(C=O), 167.89(C=O), 176.14 (C=S); MS m/z (%): 386 . Anal. Calcd. for “C19H22N4O3S” (386): C, 59.05; H, 5.74; N, 14.50; Found: C, 58.98; H, 5.72; N, 14.44%.4--6-methyl-2-thioxo-1,2,3,4-tetrahydropyrimidine-5-carbohydrazide (11): Ethyl 4--6-methyl-2-thioxo-1,2,3,4-tetrahydropyrimidine-5-carboxylate (10)  and hydrazine hydrate (5mmol) were mixed in 20 mL absolute ethanol. The reaction mixture was heated under reflux for 5hrs. After cooling, the solid formed was separated and recrystallized from ethanol as white crystals; Yield (85%); m.p. 225–227 °C, FT-IR : 3415 (NH), 3386 ,1664 (N-C=O), 1682 (C=O), 1620 (C=N), 1590 (C=C); 1H-NMR (DMSO-d6): δ 2.12 , 3.34 , 3.65 , 5.36 , 6.15–6.55 , 8.06 , 8.79 ; 13C-NMR : δ 15.72 (CH3), 17.94(CH3), 53.22 , 104.84 (Ar), 108.44 (Ar), 121.22 (Ar), 123.61(Ar), 124.92(Ar), 133.57(Ar), 143.09(Ar), 152.98(Ar), 164.58(C=O), 165.19 (C=O), 182.12 (C=S); MS m/z (%): 372. Anal. Calcd. for “C17H20N6O2S” (372): C, 54.82; H, 5.41; N, 22.56; Found: C, 54.86; H, 5.37; N, 22.51%.1,5-dimethyl-4--2-thioxo-1,2,3,4-tetrahydropyrimidin-4-yl)-2-phenyl-1,2-dihydro-3H-pyrazol-3-one (12): 4--6-methyl-2-thioxo-1,2,3,4-tetra hydropyrimidine-5-carbohydrazide (11)  and ethyl acetoacetate  were refluxed in 20 mL ethanol containing 1–3 drops of acetic acid for 5h; the solid formed while heating was separated and recrystallized from acetic acid as beige crystals. Yield: 71%, m.p. 271–273 °C, FT-IR : 3412, 3374 (NH), 1680 , 1665 (N-C = O), 1600 (C=N), 1560 (C=C); 1H-NMR (DMSO-d6): δ 1.94 , 2.26 , 3.12 , 3.34 , 3.95, 7.32–7.52 , 8.25 , 10.01 ; 13C-NMR : δ 15.72 (CH3), 17.94 (CH3), 42.15 (CH2-pyrazole), 51.22 , 106.14 (Ar), 108.24 (Ar), 120.12 (Ar), 121.61 (Ar), 122.12 (Ar), 123.01 (Ar), 134.57 (Ar), 143.09 (Ar), 152.98 (Ar), 159.14 (Ar), 163.24 (C=O), 164.58 (C=O), 165.19 (C=O), 182.12 (C=S); MS m/z (%): 438 . Anal. Calcd. for “C21H22N6O3S” (438): C, 57.52; H, 5.06; N, 19.17; Found: C, 57.56; H, 5.02; N, 19.12%.Escherichia coli ATCC 25922 and Pseudomonas aeruginosa ATCC 27853), Gram-positive bacteria (Staphylococcus aureus ATCC 25923 and Bacillus subtilis ATCC 6051), and unicellular fungi . The agar well diffusion method was used to test the antimicrobial activity of the synthesized samples against six microorganisms, including Gram-negative bacteria .The pathogenic microorganisms that were screened were known to cause UTIs. While the fungal strains were seeded on malt extract agar (MEA) plates and cultured for 3–5 days at 28 ± 2 °C before being maintained at 4 °C for subsequent usage, the examined bacteria were inoculated on nutrient agar for one day at 37 °C . The minThe specific and fixed wavelength used for detecting the optical density for the bacterial growth (at 600 nm) was completely different from the optical density for the synthesized sample. A negative control  and positive control (the tested microbes and the nutrient broth) were conducted for these purposes; additionally, another control like the synthesized sample alone was used to compare the conducted results as an auto-zero purpose. The MICs were determined as the lowest sample dilution capable of inhibiting visible bacterial growth.To evaluate the antimicrobial capability of the synthesized materials, the ZOI test must be combined with the use of nystatin (NS) as an antifungal positive control and amoxicillin/clavulanic acid , a common antibacterial drug.E. coli, the most sensitive bacteria. In 5.0 mL nutrient broth tubes, the bacterial suspension was adjusted to 0.5 McFarland (1 × 108 CFU/mL). Each tube under inspection had a single addition of compound 9. At 2 h intervals up to 24 h after treatment, the absorbance of the bacterial growth was measured (wavelength: 600 nm). To obtain the typical growth curve, the average of the triplicate data was compared to the hourly intervals [According to Huang et al., the growth curve test was used to investigate how chemical 9 affected the growth of ntervals ,52.8 CFU/mL) was chosen for the pure 18 h bacterial culture, and 100 µL of it was put into 10 mL of the nutritional broth containing compound 9. As a control, a broth devoid of component 9 was infused with culture. All of the treated samples were centrifuged for 15 min at 5000× g rpm after being incubated at 37 °C for 5 h. A total of 100 µL of supernatants and 1 mL of Bradford reagent were blended for each of the various samples. At 595 nm, optical density was measured following 10 min of incubation in the dark. Furthermore, 0.5 McFarland (1 × 108 CFU/mL) in nutrient broth tubes, and about 5.0 mL of the adjusted bacterial suspension was mixed with 1.0 mL of the prepared compound 9 at a concentration of 15.62 µg/mL  and treated microorganisms.The bacterial suspensions were adjusted to 0.5 McFarland (1 × 10p < 0.05) and Duncan’s methods. SPSS software version 15 was used to examine the accepted results. It must be noted that all the biological tests were repeated three times.The statistical analysis of the obtained results was investigated after applying a one-way ANOVA (at https://www.rcsb.org/) [E. coli β-carbonic anhydrase [The 2D structures of compounds were sketched using ChemDraw 16 and then converted to 3D structures and minimized using OpenBabel GUI . The tarsb.org/)  under PDnhydrase . The filnhydrase  in Disconhydrase . The dugE. coli, P. aeruginosa, S. aureus, B. subtilis, C. albicans, and C. neoformans, whereas the MICs for compounds 9, 5, and 4 were 15.62, 31.25, and 125 µg/mL, respectively. It was discovered that the amount of cellular protein released from E. coli is directly correlated to the quantity of compound 9, which was found to be 177.99 µg/mL following treatment with 1.0 mg/mL of compound 9. This finding supports compound 9’s antibacterial properties and explains how the formation of pits in the E. coli cell membrane results in the release of proteins from the cytoplasm. A substance must stick to its target site on microbial cells and establish a specific number of crucial regions associated with its level within pathogenic microorganisms in order for it to possess antimicrobial potentials that may eliminate those pathogenic microbes. The docking studies proved that compound 9 declared a greater activity than other synthesized compounds.The present study overlays the synthesis of a new series of thiadiazole and pyrimidine moieties attached to antipyrine analogue 2–12, and their structures were confirmed via spectral and elemental analyses. The antimicrobial potential was estimated through in vitro ZOI and MIC. On the other hand, the antibacterial reaction mechanism was tested through a membrane leakage assay and SEM imaging technique. In comparison to AMC/Nyst, the industry standard antimicrobial agent, all developed compounds showed promising antibacterial activity against all of the evaluated bacterial and fungal species. In comparison to AMC/Nyst, compounds 4, 5, and 9 were noticeably more active. With inhibition zones of 19.0, 14.2, 17.0, 15.0, 16.9, and 9.5 mm, respectively, compound 9 at a dosage of 0.1 mg/mL showed potential antibacterial action against


Palladium-catalyzed arylthioallylation reactions then allowed the introduction of 3′-arylthiol substituents. We also report for the first time the synthesis of the 4-methoxybenzyl ester of -3-[(acetyloxy)methyl]-7-amino-7-methoxy-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid and the use of diphenyl trichloroacetimidate, instead of diphenyldiazomethane, and 4-methoxybenzyl trichloroacetimidate to prepare related 4-methoxybenzyl esters. The chemistry described, and several of the synthetic intermediates reported here, are potentially valuable methods and scaffolds, respectively, for further development of β-lactam antibiotics. The aim of this project was to develop a synthetic protocol for the preparation of a cephamycin scaffold that would readily allow the synthesis of its analogues with variations at the C-7 amino group and the C-3′ position. We also aimed to develop a method that avoided the use of toxic and potentially explosive diphenyldiazomethane. These aims were achieved via the synthesis of the novel α-bromo acetamide  The latter two compounds are biosynthetically connected to penicillin N 1 by ring-expansion and, in the case of cephamycin C 3, enzymatic introduction of a 7-α-methoxy substituent . The spores are commonly associated with transmission, relapse, and recurrence of CDI and inhibiting this sporulation process could potentially prevent the recurrence of CDI [4 we required an advanced synthetic intermediate that would allow the synthesis of analogues (10) with variations at the C-7 amino group and the C-3′ position for structure–activity relationships studies and future antibacterial drug development -3-[(acetyloxy)methyl]-7-amino-7-methoxy-8-oxo-5-thia-1-azabicyclo [4.2.0]oct-2-ene-2-carboxylic acid] would have been the ideal precursor it was prohibitively expensive  ..4–6 when7 from 7-ACA 9 that avoids the use of toxic and potentially explosive diphenyldiazomethane bis (BIPHEPHOS) as a ligand and acetonitrile as a solvent -oct-2-ene-2-carboxylic acid 12 in anhydrous DMA (5 mL) under N2 at rt, BSA  was added and stirred for 30 min until the mixture turned clear. The reaction mixture was then cooled to −20 °C (NaCl/ice), and phenylacetyl chloride  was added dropwise. The resulting solution was stirred at −20 °C for 2 h, at which point the reaction was shown to be complete via TLC analysis (TLC (MeOH/CH2Cl2—2:3): Rf = 0.63). The reaction mixture was poured into iced water (20 mL) and extracted using EtOAc (3 × 20 mL). The combined EtOAc layer was washed with water (3 × 20 mL), brine (20 mL), dried over anhydrous MgSO4, filtered, and concentrated in vacuo to give a pale-yellow residue. The obtained crude product was then dissolved in a minimum amount of EtOAc, and hexanes were added dropwise to this vigorously stirred solution until precipitation started to occur. The mixture was stirred at rt overnight. The solvent was removed using a syringe and the precipitate was washed with hexanes (5 × 10 mL), then dried in vacuo to afford the titled compound as an off-white powder . 1H NMR  δ 7.95 , 7.26–7.07 , 5.71 , 5.00 , 4.83 , 3.58–3.47 , 3.39 , 1.90 ; the 1H NMR spectroscopic data were in agreement with those previously reported oct-2-ene-2-carboxylate 13 oct-2-ene-2-carboxylic acid 14 in DMA (1 mL) BSA  was added, and the reaction mixture was stirred at rt for 30 min until the white suspension turned into an orange solution. Powdered molecular sieves  and 3,5-di-tert-butyl-4-hydroxybenzaldehyde  were then added to the reaction mixture and stirred at rt for 2 h until the pale-orange color of the reaction turned into bright yellow. The completion of the reaction was indicated via TLC analysis (TLC (MeOH/CH2Cl2—2:3): Rf = 0.76). A portion of the reaction mixture was diluted with MeOH (10 mL) and filtered. The obtained filtrate was concentrated for NMR analysis, which indicated a mixture of the titled compound and 3,5-di-tert-butyl-4-hydroxybenzaldehyde. The reaction mixture was concentrated under a stream of N2 overnight to give crude product 14 as a yellow gum. 1H NMR  δ 8.44 , 7.57 , 5.53 , 5.28 , 4.84 , 3.53 , 2.04 , 1.39 3); (ESI +ve) m/z 489 .To a suspension of 7-ACA R,7R)-3-(acetoxymethyl)-7-(((E)-3,5-di-tert-butyl-4-hydroxy-benzylidene)amino)-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylate 15aBenzhydryl  in CH2Cl2 (2 mL) under an Ar atmosphere at rt benzhydryl 2,2,2-trichloroacetimidate  was added. The color of the reaction turned from bright yellow to brown, and the completion of the reaction was indicated using TLC analysis (TLC (MeOH/CH2Cl2—1:9): Rf = 0.74). The reaction mixture was concentrated in vacuo and the obtained brown residue was suspended in MeOH (30 mL) and filtered. The filtrate was concentrated for NMR analysis, which indicated a mixture of the titled compound, DMA and 3,5-di-tert-butyl-4-hydroxybenzaldehyde. Further purification using flash chromatography resulted in decomposition. 1H NMR  δ 8.45 , 7.65 , 7.46–7.22 , 6.94 , 5.46 , 5.24 , 4.98 , 4.71 , 3.56 , 1.98 , 1.46 3); the 1H NMR spectroscopic data agreed with those previously reported oct-2-ene-2-carboxylate 16aBenzhydryl  in MeOH (1.5 mL) at −20 °C DDQ  was added, and the reaction mixture was stirred at −20 °C for 45 min. The completion of the reaction was indicated using MS analysis. EtOAc (1.5 mL) was then added to the reaction and the resulting mixture was stirred for another 30 min. The dark red reaction mixture was concentrated in vacuo and purified via flash chromatography over SiO2 (EtOAc/hexane—1:10–3:10) to give the title compound as a hydroscopic, yellow foam . To a suspension of crude product 9). Alternative purification method for scale-up reactions  δ 8.54 , 7.69 , 7.51–7.29 , 6.97 , 5.64 , 5.08 , 4.85 , 3.57 , 3.39 , 2.00 , 1.46 3); the 1H NMR spectroscopic data agreed with those previously reported oct-2-ene- 2-carboxylate 7Benzhydryl  in EtOAc (0.5 mL) a solution of Girard-T reagent  was added in MeOH (0.6 mL) at rt, and the resulting solution was stirred for 2.5 h until the completion of the reaction was indicated by MS analysis. Upon completion, the reaction mixture was diluted with EtOAc (15 mL) and poured into water (20 mL), which was extracted with additional EtOAc (15 mL × 2). The combined EtOAc layer was washed with brine (20 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo to give the crude product as a dark green gum. Further purification of the crude product resulted in decomposition and it was, therefore, used directly for the subsequent reaction. 1H NMR  δ 7.41–7.27 , 6.94 , 5.03 , 4.85 , 4.83 , 3.52 , 3.44 , 3.31 , 2.02 ; (ESI +ve) m/z 469 , 491 , and 501 . The sample of 7 also contained (E)-2-hydrazineyl)-N,N,N-trimethyl-2-oxoethan-1-aminium chloride in a 1:1 ratio from 1H NMR analysis: 1H NMR  δ 8.38 , 7.57 , 4.65 , 3.42 , 1.43 . To a solution of R,7S)-3-(acetoxymethyl)-7-(2-bromoacetamido)-7-methoxy-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylate 17aBenzhydryl  under N2 at −20 °C (NaCl/ice) pyridine  was added and the resulting solution was stirred for 2 min. Bromoacetyl bromide  was then added dropwise to the solution. The reaction mixture was stirred at −20 °C for 2 h, at which point the reaction was shown to be complete via TLC analysis (TLC (EtOAc/hexane—2:3): Rf = 0.39). The reaction mixture was poured into EtOAc (5 mL); washed sequentially with HCl (1.0 M—5 × 5 mL), saturated NaHCO3 (5 × 5 mL), and brine (10 mL); dried over anhydrous Na2SO4; filtered; and concentrated in vacuo to give a brown residue. The crude product was purified via flash chromatography over SiO2 (EtOAc/hexane—1:19–9:11) to give the titled compound as a pale-yellow foam . c 0.22, CH2Cl2); 1H NMR  δ 7.51–7.23 , 6.95 , 5.09 , 4.95 , 3.92 , 3.58 , 3.39 , 2.02 ; NH signal was not observed; 13C NMR  δ 170.4 (C4′), 166.6 , 160.2 (C2ʹ), 160.1 (C8), 139.3 , 139.1 , 132.0 (C3), 128.57 (ArCH), 128.55 (ArCH), 128.5 (ArCH), 128.2 (ArCH), 128.2 (ArCH), 127.5 (ArCH), 127.04 (ArCH), 127.01(ArCH), 125.9 (C2), 95.8 (C7), 79.7(CH-Ph2), 64.3 (C6), 62.4 (C3′), 54.0 (O-CH3), 27.9 (Br-CH2), 27.0 (C4), 20.64 (CH3); IR (cm−1) max 3278 (w), 2960 (w), 1775 , 1735 , 1608 (m), 1517 (s), 1380 (m), 1237 , 1129 (m), 1090 (m), 1032 (m), 749 (w), 700 (m); MS (ESI + ve) m/z 611 , 613 ; HRMS (ESI +ve TOF) calcd for 79BrC26H25N2O7SNa 611.0464, found 611.0435 ([M + Na]+).To a solution of crude product R,7R)-3-(acetoxymethyl)-7-(((E)-3,5-di-tert-butyl-4-hydroxy-benzylidene)amino)-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylate 15b4-Methoxybenzyl  in CH2Cl2 (2 mL) under an Ar atmosphere at rt 4-methoxybenzyl 2,2,2-trichloroacetimidate  was added, and the resulting mixture was stirred at rt for 48 h. The color of the reaction turned from bright yellow to brown, and the completion of the reaction was indicated using MS analysis. The reaction mixture was filtered and concentrated in vacuo to give the crude product 15 as a sticky brown gum. A portion of this filtrate was used for NMR analysis, which indicated a mixture of the titled compound, DMA, and 3,5-di-tert-butyl-4-hydroxybenzaldehyde. Further purification using flash chromatography resulted in decomposition. 1H NMR  δ 8.42 , 7.63 , 7.35 , 6.90 , 5.40 , 5.25–5.15 , 4.99 , 4.76 , 3.78 , 3.64 , 3.46 , 2.02 , 1.44 3); (ESI + ve) m/z 609  and 631; (ESI − ve) m/z 607 .].To a suspension of crude product R,7S)-3-(acetoxymethyl)-7-(2-bromoacetamido)-7-methoxy-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylate 16bBenzhydryl (615b was suspended in MeOH (5 mL) at −20 °C under an Ar atmosphere. To the suspension DDQ  was added, and the reaction mixture was stirred at −20 °C for 45 min. The completion of the reaction was indicated via TLC analysis (TLC (EtOAc/hexane—2:3): Rf = 0.39). The reaction mixture was concentrated in vacuo and purified via column chromatography over SiO2. Prior to the purification, a series of solvents in a EtOAc/hexane system were prepared  and chilled at −20 °C overnight. On the following day, an appropriately sized column was packed with a SiO2 slurry in hexane and chilled at −20 °C until ready to use. Once the crude product was loaded onto the column, the pre-chilled solvents was kept cool on an ice bath and the flow rate was accelerated by using a stream of compressed air to ensure the entire purification process was kept within 30 min. Fractions with Rf value of 0.39 (TLC (EtOAc/hexane—2:3)) were combined and concentrated in vacuo to give the title compound as a hydroscopic, pale-yellow foam . c 0.27, CH2Cl2); 1H NMR  δ 8.51 , 7.69 , 7.39 , 6.90 , 5.63 , 5.27 , 5.05 , 4.87 , 3.81 , 3.55 , 3.38 , 2.03 , 1.46 3); 13C NMR  δ 170.6 (C4′), 165.8 (C9), 163.4 (C8), 161.5 (C2′), 160.0 , 157.9 (C-OH), 136.3 (C-C(CH3)3), 130.7 , 126.9 (C10), 126.5 (C-C9), 126.2 , 124.2 (C2), 114.0 (C12), 104.6 (C7), 68.1 , 64.5 (C6), 63.1 (C3′), 55.3 , 53.5 , 34.4 (C(CH3)3), 30.2 (C(CH3)3), 26.6 (C4), 20.7 ; C3 resonance was not observed in the 13C NMR or HMBC spectra; IR (cm−1) max 3613 (w), 2958 (m), 1772 , 1737 , 1631 (m), 1516 (m), 1429 (m), 1389 (m), 1302 (w), 1235 , 1176 (m), 1129 (m), 1093 (m), 1034 (m), 888 (w), 827 (m), 775 (w); MS (ESI +ve) m/z 639 , 661 , 1300 ; (ESI − ve) m/z 637 ; HRMS (ESI + ve TOF) calcd for C34H43N2O8S 639.2740, found 639.2752 ([M + H]+).The crude compound R,7S)-3-(acetoxymethyl)-7-amino-7-methoxy-8-oxo-5-thia-1 azabicyclo[4.2.0]oct-2-ene-2-carboxylate 84-Methoxybenzyl  in EtOAc (2.4 mL) a solution of Girard-T reagent  in MeOH (2.9 mL) at rt was added, and the resulting solution was stirred for 3.5 h until the completion of the reaction was indicated via TLC analysis (TLC (EtOAc/hexane—1:1): Rf = 0.24). Upon completion, the reaction mixture was diluted with EtOAc (50 mL) and poured into water (50 mL), which was extracted using additional EtOAc (50 mL × 2). The combined EtOAc layer was washed with brine (70 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo to give the crude product as a dark green gum. A portion of this crude product (0.216 mmol) was purified via flash chromatography over SiO2 (EtOAc/hexane—1:10–2:3) to give the titled compound as a green gum . c 0.77, CH2Cl2); 1H NMR  δ 7.35 , 6.88 , 5.23 , 5.02 , 4.85–4.76 , 3.80 , 3.53–3.25 , 2.04 ; 13C NMR  δ 170.8 (C4′), 164.1 (C8), 161.5 (C2′), 160.1 , 130.8 , 127.1 , 126.7 (C2), 114.1 , 98.6 (C7), 68.2 , 63.9 (C6), 63.0 (C3′), 55.5 , 52.6 , 26.8 (C4), 20.9 ; C3 resonance was not observed in the 13C NMR or HMBC spectra; IR (cm−1) max 3307 (w), 2956 (w), 2930 (w), 1777 , 1725 , 1612 (m), 1514 (s), 1455 (w), 1379 (m), 1354 (m), 1219 , 1174 (s), 1112 (m), 1026 (s), 821 (s), 728 (m); MS (ESI + ve) m/z; 445 , 468 , 867 ; HRMS (ESI + ve TOF) calcd for C19H22N2O7SNa 455.1045, found 455.1063 ([M + Na]+).To a solution of R,7S)-3-(acetoxymethyl)-7-(2-bromoacetamido)-7-methoxy-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylate 17b4-Methoxybenzyl  in anhydrous CH2Cl2 (3 mL) under N2 at –20 °C (NaCl/ice) pyridine  was added and the resulting solution was stirred for 2 min. Bromoacetyl bromide  was then added dropwise to the solution. The reaction mixture was stirred at –20 °C for 2 h, at which point the reaction was shown to be complete via TLC analysis (TLC (EtOAc/hexane—2:3): Rf = 0.32). The reaction mixture was poured into EtOAc (5 mL); washed sequentially with HCl (1.0 M—5 × 50 mL), saturated NaHCO3 (5 × 50 mL), and brine (100 mL); dried over anhydrous Na2SO4, filtered, and concentrated in vacuo to give a brown residue. The crude product was purified via flash chromatography over SiO2 (EtOAc/hexane—1:19–9:11) to give the titled compound as an orange gum . c 1.08, CH2Cl2); 1H NMR  δ 7.36 , 6.89 , 5.25 , 5.07 , 4.94 , 3.93 , 3.81 , 3.56 , 3.40 , 2.05 , NH signal was not observed; 13C NMR  δ 170.5 (C4′), 166.7 , 160.9 (C2′), 160.1 (C8), 159.9 , 130.6 , 126.8 , 126.0 (C2), 114.0 (C12), 95.6 (C7), 68.1 , 62.6 (C3′), 55.3 , 53.9 , 27.9 (Br-CH2), 26.9 (C4), 20.7 ; C3 resonance was not observed in the 13C NMR or HMBC spectra; IR (cm−1) max 3279 (w), 2962 (w), 2838 (w), 1777 , 1735 , 1613 (m), 1516 (s), 1392 (m), 1240 , 1178 (m), 1130 (m), 1088 (m), 10320 (m), 853 (w); MS (ESI + ve) m/z 565 , 567 , 588 , 590 ; HRMS (ESI + ve TOF) calcd for 79BrC21H23N2O8SNa 565.0256, found 565.0266 ([M + Na]+).To a solution of the crude compound R,7S)-3-(Acetoxymethyl)-7-(2-bromoacetamido)-7-methoxy-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid 18 in anisole  TFA was added , and the solution was stirred at rt for 10 min. The reaction mixture was poured into EtOAc (50 mL) and extracted with saturated NaHCO3 (3 × 50 mL). The combined aqueous layer was washed with EtOAc (2 × 50 mL). Additional EtOAc (200 mL) was added to a stirred solution of the aqueous layer, and the aqueous layer was acidified with conc. HCl to pH < 1. The two layers were then separated. The aqueous layer was extracted with EtOAc (2 × 50 mL), and the combined organic layer was washed sequentially with HCl (1.0 M—3 × 50 mL) and brine (50 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo to give a yellow gum. This crude product was redissolved in a minimum amount of EtOAc, and hexane was added dropwise to the vigorously stirred solution of EtOAc until precipitation started to occur. The mixture was stirred at rt overnight. The solvent was removed using a syringe and the precipitate was washed with hexanes (5 × 10 mL) then dried in vacuo to afford the titled compound as a yellow gum . Method 1—prepared from compound 17b. To a solution of compound 17b  in anhydrous CH2Cl2 (1 mL) at 0 °C TFA was slowly added . The resulting dark-pink solution was stirred at 0 °C for 30 min. Work-up and precipitation as in Method 1 gave the titled compound as a yellow gum . c 1.41, MeOH); 1H NMR  δ 7.79 , 5.06 , 4.87 , 3.89 , 3.56 , 3.50 , 3.33 , 2.02 ; 13C NMR  δ 171.1 (C4′), 167.9 , 162.4 (C2′), 160.9 (C8), 128.7 (C3), 126.1 (C2), 95.6 (C7), 64.2 (C6), 63.0 (C3′), 53.6 (O-CH3), 28.5 (Br-CH2), 26.6 (C4), 20.5 ; IR (cm−1) max 3538 (w), 3271 (w), 3022 (w), 1774 ,1728 , 1693 (s), 1539 (m), 1386 (m), 1235 , 1134 (m), 1088 (m), 1024 (m); MS (ESI + ve) m/z 445 , 447 ; HRMS (ESI + ve TOF) calcd for 79BrC13H15N2O7S2Na 444.9681, found 444.9699 ([M + Na]+).Method 2—prepared from compound R,7S)-3-(Acetoxymethyl)-7-(2-azidoacetamido)-7-methoxy-8-oxo-5-thia-1-azabicyclo [4.2.0]oct-2-ene-2-carboxylic acid 19 in DMF (0.3 mL) at −15 °C sodium azide  was added, and the solution was stirred for 24 h, at which point the reaction was shown to be complete via MS analysis. The reaction mixture was diluted with distilled H2O (20 mL), to which EtOAc (20 mL) was added. The resulting mixture was stirred vigorously, and the aqueous layer was acidified with conc. HCl to pH < 1. The two layers were separated, and the aqueous layer was extracted with EtOAc (3 × 20 mL). The combined organic layer was washed sequentially with distilled H2O (3 × 20 mL) and brine (40 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo to give a sticky gum. This crude product was dissolved in a minimum amount of EtOAc, and hexane was added dropwise to the vigorously stirred solution until the precipitation started to occur. The mixture was stirred at rt overnight. The solvent was removed using a syringe and the precipitate was washed with hexanes (5 × 10 mL), then dried in vacuo to afford the titled compound as a pale-yellow gum . c 0.77, MeOH); 1H NMR  δ 7.58 , 5.06 , 4.88 , 3.95 , 3.57 , 3.50 , 3.34 , 2.02 ; 13C NMR  δ 171.1 (C4′), 169.2 , 162.5 (C2′), 161.0 (C8), 128.6 (C3), 126.3 (C2),96.3 (C7), 64.1 (C6), 63.0 (C3′), 53.6 (O-CH3), 51.6 (N3-CH2), 26.7 (C4), 20.5 ; IR (cm−1) max 3385 (m), 3223 (m), 3026 (w), 2111 (N=N=N stretching), 1772 , 1705 (s) 1514 (m), 1424 (m), 1385 (m), 1230 , 1134 (m), 1087 (m), 1026 (m), 551 (w); MS (ESI +ve) m/z 403 , 408 , 431 , 449 ; (ESI − ve) m/z 384 , 420 , 422 , 498 ; HRMS (ESI + ve TOF) calcd for C13H15N5O7S2Na 408.0590, found 408.0581 ([M + Na]+).To a solution of compound 20 and 21a–h, the following numbering system has been used.For NMR assignments of compounds R,7S)-3-(Acetoxymethyl)-7-methoxy-8-oxo-7-acetamido)-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid 20, CuSO4·5H2O  and sodium ascorbate  a mixture of t-BuOH and H2O  was added. To this stirred mixture, phenylacetylene  was then added, and the reaction mixture was stirred at 30 ℃ for 24 h. The reaction mixture was diluted with EtOAc (20 mL), washed with saturated aqueous NH4Cl solution (20 mL), dried over anhydrous MgSO4, filtered, and concentrated in vacuo. The obtained residue was then dissolved in a minimum amount of EtOAc, and to this vigorously stirred solution hexanes were added dropwise until precipitation started to occur. The mixture was stirred at rt overnight. The solvent was removed using a syringe and the precipitate was washed with hexanes (5 × 10 mL), then dried in vacuo to afford the titled compound as a thin, yellow film . c 0.54, MeOH); 1H NMR  δ 8.16 , 7.93 , 7.89–7.84 , 7.46 , 7.39–7.33 , 5.28 , 5.06 , 4.88 , 3.59–3.50 , 3.32 , 2.02 ; 13C NMR  δ 171.1 (C4′), 167.2 , 162.6 (C2′), 160.9 (C8), 147.7 , 131.3 (C1″), 129.5 (C3″), 128.71 (C3) 128.68 (C4″), 126.3 (C2), 126.1 (C2″), 122.9 , 96.4 (C7), 64.1 (C6), 63.0 (C3′), 53.8 (O-CH3), 52.4 (CH2-triazole), 26.7 (C4), 20.5 ; IR (cm−1) max 3288 (w), 2946 (w), 1779 , 1708 (s), 1533 (m), 1442 (w), 1382 (m), 1231 , 1136 (m), 1087 (m) 1028 (m), 768 (s), 696 (m); MS (ESI + ve) m/z 488 , 510 , 975 ; (ESI − ve) m/z 486 ; HRMS (ESI + ve TOF) calcd for C17H17N5O7SNa 510.1059, found 510.1050 ([M + Na]+).To a reaction vessel charged consecutively with azide 21a–h. Synthesis of the thiomethyl compound 21a is given as an example. The General Procedure for the Pd/BIPHEPHOS reaction for the preparation of the cephamycin C-3 thiol derivatives R,7S)-7-Methoxy-3-(((4-methoxyphenyl)thio)methyl)-8-oxo-7-acetamido)-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid 21a(62(dba)3·CHCl3 , BIPHEPHOS , and anhydrous MeCN (2 mL) were added under Ar at 50 °C. The resulting suspension was stirred at 50 °C for 30 min until it turned into a bright-yellow solution. The reaction flask was allowed to cool to rt and was then placed in a sonicator with the water temperature at 30–35 °C. Compound 20  and 4-methoxybenzenethiol  were added to the reaction flask. The resulting mixture was sonicated at 30–35 °C until the completion of the reaction (10 h) was indicated using MS analysis. The reaction mixture was poured into EtOAc (10 mL) and extracted with NaHCO3 (3 × 10 mL). The combined aqueous layer was acidified with conc. HCl to pH = 3 and extracted with EtOAc (3 × 10 mL). The combined EtOAc fraction was then washed with HCl (1.0 M—3 × 10 mL) and brine (20 mL), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo to give a yellow gum. This crude product was purified via semi-preparative RP-HPLC using a MeCN/H2O gradient mobile phase containing 0.01% TFA  at a flow rate of 3.8 mL/min to give the titled compound as a pale-yellow oil . Compound purity by HPLC ; 1H NMR  δ 8.16 , 7.87 , 7.45 , 7.42–7.33 , 6.85 , 5.28 , 4.97 , 4.21 , 3.77 , 3.71 , 3.55 , 3.53 , 3.35 ; NH resonance was not observed; 13C NMR  δ 167.2 , 162.5 (C2′), 161.2 (C8), 160.9 (C8″), 148.0 , 137.3 (C3), 136.7 (C6″), 131.6 (C1″), 129.7 (C3″), 128.9 (C4″), 126.3 (C2″), 125.3 (C2), 124.4 (C5″), 123.1 , 115.4 (C7″), 97.0 (C7), 65.9 (C6), 55.9 , 54.0 (Ar-OMe), 52.8 (CH2-triazole), 38.8 (C3′), 29.48 (C4); IR (cm−1) max 3267 (w), 2993 (w), 2839 (w), 1771 , 1703 (s), 1591 (m), 1494 (s), 1465 (w), 1442 (w), 1286 (w), 1247 (s), 1181 (w), 1148 (w), 1106 (m), 1088 (m), 1026 (m), 830 (m), 766 (s), 696 (m); MS (ESI + ve) m/z 568 , 590 ; (ESI − ve) m/z 566 , 680 ; HRMS (ESI − ve TOF) calcd for C26H24N5O6S2 566.1168, found 566.1160 ([M − H]−).This compound was prepared following a procedure reported in the literature with some modifications . To a flHPLC see : 99.8%, R,7S)-3-(((4-Carboxyphenyl)thio)methyl)-7-methoxy-8-oxo-7-acetamido)-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid 21b, Pd2(dba)3·CHCl3 , BIPHEPHOS , and 4-mercaptobenzoic acid  in MeCN (2 mL) with 19 h of reaction time. Work-up and purification via RP-HPLC  as described above gave the titled compound as a pale-yellow oil . Compound purity by HPLC ; 1H NMR  δ 8.14 , 7.99–7.82 , 7.80 , 7.54–7.40 , 7.40–7.31 , 5.26 , 4.98 , 4.31 , 4.01 , 3.54 , 3.53 , 3.36 ; 13C NMR  δ 167.1 , 167.0 , 162.5 (C2′), 161.0 (C8), 147.8 , 142.0 (C5″), 135.1 (C3), 131.3 (C1″), 130.7 (C3″), 130.5 (C7″), 129.5 (C6″), 128.9 (C4″), 128.7 (C8″), 126.1 (C2″), 125.5 (C2), 122.9 , 96.7 (C7), 65.4 (C6), 53.8 (O-CH3), 52.5 (CH2-triazole), 35.9 (C3′), 29.0 (C4); IR (cm−1) max 3527 (w), 3032 (w), 2940 (w), 1772 , 1701 (s), 1592 (m), 1560 (w), 1402 (w), 1364 (w), 1235 (s), 1182 (w), 1110 (m), 1187 (m), 1015 (w), 851 (w), 796 (w), 765 (s), 695 (m); MS (ESI + ve) m/z 582 , 604 ; (ESI − ve) m/z 580 , 680 ; HRMS (ESI − ve TOF) calcd for C26H22N5O6S2 580.0961, found 580.0952 ([M − H]−).This compound was prepared according to the General Procedure using HPLC see : 98.7%, R,7S)-3-(((4-Cyanophenyl)thio)methyl)-7-methoxy-8-oxo-7-acetamido)-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid 21c, Pd2(dba)3·CHCl3 , BIPHEPHOS  and 4-mercaptobenzonitrile  in MeCN (2 mL) with 13 h of reaction time. Work-up and purification via RP-HPLC  as described above gave the titled compound as a thin, transparent film . Compound purity by HPLC ; 1H NMR  δ 8.14 , 7.86 , 7.81 , 7.61 , 7.46 , 7.40–7.31 , 5.26 , 4.98 , 4.29 , 4.05 , 3.53 , 3.52 , 3.35 ; 13C NMR  δ 167.0 , 161.0 (C8), 147.8 , 142.8 (C5″), 134.0 (C3), 133.1 (C7″), 131.4 (C1″), 130.6 (C3″), 129.5 (C6″), 128.7 (C4″), 126.1 (C2″), 125.4 (C2), 122.9 , 119.1 (C8″), 110.2 (C≡N), 96.6 (C7), 65.4 (C6), 53.8 (O-CH3), 52.5 (CH2-triazole), 35.6 (C3′), 28.9 (C4); C2′ resonance was not observed in the 13C NMR or HMBC spectra; IR (cm−1) max 3500 (w), 3279 (w), 2228 , 1771 , 1705, 1592 (m), 1537 (w), 1468 (w), 1372 (m), 1154 (w), 1017 (s), 827 (m), 767 (s), 696 (m), 549 (m); MS (ESI + ve) m/z 585 , 607 , 639 ; (ESI − ve) m/z 561 , 675 ; HRMS (ESI +ve TOF) calcd for C26H23N6O5S2 563.1171, found 563.1166 ([M + H]+).This compound was prepared according to the General Procedure using HPLC see : 99.9%, R,7S)-3-(((4-Fluorophenyl)thio)methyl)-7-methoxy-8-oxo-7-acetamido)-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid 21d, Pd2(dba)3·CHCl3 , BIPHEPHOS , and 4-fluorobenzenethiol  in MeCN (2 mL) with 23 h of reaction time. Work-up and purification via RP-HPLC  as described above gave the titled compound as a pale-yellow oil . Compound purity by HPLC ; 1H NMR  δ 8.15 , 7.86 , 7.49–7.42 , 7.36 , 7.05 , 5.27 , 4.97 , 4.24 , 3.79 , 3.53 , 3.52 , 3.35 , NH resonance was not observed; 13C NMR  δ 167.0 , 163.3 , 162.5 (C2′), 161.0 (C8), 147.8 , 136.5 , 136.1 (C3), 131.4 (C1″), 129.5 (C3″), 128.7 (C4″), 126.1 (C2″), 125.5 (C2), 122.9 , 116.55 , 96.7 (C7), 65.7 (C6), 53.8 (O-CH3), 52.5 (CH2-triazole), 38.1 (C3′), 29.1 (C4), C5″ resonance was not observed in the 13C NMR or HMBC spectra; IR (cm−1) max 3283 (w), 3144 (w), 2942 (w), 1768 , 1699 (s), 1589 (m), 1534 (m), 1490 (s), 1370 (m), 1220 , 1156 (m), 1109 (m), 1088 (m), 1016 (w), 832 , 766 (s), 695 (m), 629 (w), 517 (w); MS (ESI +ve) m/z 556 , 578 ; (ESI − ve) m/z 554 , 668 ; HRMS (ESI − ve TOF) calcd for C25H21N5O5S2F 554.0968, found 554.0976 ([M − H]−).This compound was prepared according to the General Procedure using HPLC see : 99.6%, R,7S)-7-Methoxy-8-oxo-7-acetamido)-3-((p-tolylthio)methyl)-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid 21e, Pd2(dba)3·CHCl3 , BIPHEPHOS , and 4-methylbenzenethiol  in MeCN (2 mL) with 13 h of reaction time. Work-up and purification via RP-HPLC  as described above gave the titled compound as a pale-yellow oil . Compound purity by HPLC ; 1H NMR  δ 8.15 , 7.89–7.84 , 7.81 , 7.45 , 7.36 , 7.31 , 7.12 , 5.27 , 4.96 , 4.22 , 3.81 , 3.52 , 3.51 , 3.33 , 2.30 ; 13C NMR  δ 167.0 , 162.4 (C2′), 161.0 (C8), 147.8 , 138.7 (C8″), 136.5 (C3), 133.5 (C6″), 131.4 (C1″), 130.8 (C5″), 130.3 (C7″), 129.5 (C3″), 128.7 (C4″), 126.1 (C2″), 125.1 (C2), 122.9 , 96.7 (C7), 65.6 (C6), 53.8 (O-CH3), 52.5 (CH2-triazole), 37.7 (C3′), 29.1 (C4), 20.8 (CH3); IR (cm−1) max 3519 (w), 3282 (w), 3024 (w), 1770 , 1703 (s), 1536 (m), 1492 (w), 1442 (w), 1373 (m), 1233 (m), 1153 (w), 1088 (m), 1018 (m), 810 (m), 766 (s), 695 (m), 503 (w); MS (ESI + ve) m/z 552 , 574 ; (ESI − ve) m/z 550 , 664 ; HRMS (ESI − ve TOF) calcd for C26H24N5O5S2 550.1219, found 550.1227 ([M − H]−).This compound was prepared according to the General Procedure using HPLC see : 99.9%, R,7S)-7-Methoxy-3-(((4-(methylthio)phenyl)thio)methyl)-8-oxo-7-acetamido)-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid 21f, Pd2(dba)3·CHCl3 , BIPHEPHOS , and 4-(methylsulfanyl)thiophenol  in MeCN (2 mL) with 13 h of reaction time. Work-up and purification via RP-HPLC  as described above gave the titled compound as a pale-yellow oil . Compound purity by HPLC ; 1H NMR  δ 8.15 , 7.89–7.83 , 7.78 , 7.45 , 7.39–7.31 , 7.18 , 5.27 , 4.97 , 4.23 , 3.81 , 3.53 , 3.52  3.35 , 2.45 ; 13C NMR  δ 167.0 , 162.4 (C2′), 161.0 (C8), 147.8 , 139.7 (C8″), 136.4 (C3), 134.2 (H6″), 131.4 (C1″), 130.2 (C5″), 129.5 (C3″), 128.7 (C4″), 127.0 (C7″), 126.1 (C2″), 125.2 (C2), 122.9 , 96.7 (C7), 65.6 (C6), 53.8 (O-CH3), 52.5 (CH2-triazole), 37.8 (C3′), 29.2 (C4), 15.1 (CH3); IR (cm−1) max 3507 (w), 3278 (w), 3139 (w), 1770 , 1704 (s), 1625 (w), 1532 (m), 1478 (w), 1440 (w), 1370 (m), 1230 (s), 1152 (s), 1106 (w), 1087 (s), 1013 (w), 1000 (w), 812 (m), 766 (s), 695 (m), 504 (w); MS (ESI + ve) m/z 584 , 606 , 622 ; (ESI −ve) m/z 582 , 696 ; HRMS (ESI − ve TOF) calcd for C26H24N5O5S3 582.0940, found 582.0931 ([M − H]−).This compound was prepared according to the General Procedure using HPLC see : 99.8%, R,7S)-3-(((4-Ethylphenyl)thio)methyl)-7-methoxy-8-oxo-7-acetamido)-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid 21g, Pd2(dba)3·CHCl3 , BIPHEPHOS , and 4-ethylbenzenethiol  in MeCN (2 mL) with 10 h of reaction time. Work-up and purification via RP-HPLC  as described above gave the titled compound as a pale-yellow oil . Compound purity by HPLC ; 1H NMR  δ 8.15 , 7.86 , 7.78 , 7.45 , 7.40–7.30 , 7.28–7.18 , 7.15 , 5.26 , 4.96 , 4.93 , 4.22 , 4.16 , 3.88 , 3.82 , 3.52 , 3.51 , 3.35 , 3.34 , 2.79 , 2.61 , 1.18 , 1.18 ; 13C NMR  δ 167.0 , 162.4 (C2′), 161.0 (C8), 147.8 , 145.0 (C8″), 136.5 (C3), 133.7 (*C6″), 133.5 (C6″), 131.4 (C5″), 131.1 (*C5″), 129.6 (*C7″), 129.5 (C3″), 129.2 (C7″), 128.7 (C4″), 126.1(C2″), 125.1 (C2), 122.9 , 96.7 (C7), 65.6 (C6), 65.4 (*C6), 53.8 (O-CH3), 52.5 (CH2-triazole), 37.6 (C3′), 37.0 (*C3′), 29.2 (C4), 28.8 (*C4), 28.7 (C9″), 27.4 (*C9″), 15.5 (CH3), 15.2 (*CH3); C1″ resonance was not observed in the 13C NMR or HMBC spectra (*resonance of minor rotamer observed in the 1H NMR and/or 13C NMR spectra); IR (cm−1) max 3280 (w), 2965 (w), 1772 , 1704 (s), 1632 (w), 1532 (m), 1466 (w), 1441 (w), 1371 (m), 1232 (m), 1153 (w), 1109 (m), 1088 (m) 1000 (w), 829 (m), 765 (s), 627 (m), 520 (w); MS (ESI + ve) m/z 566 , 588 ; (ESI − ve) m/z 564 , 600 , 628 ; HRMS (ESI + ve TOF) calcd for C27H27N5O5S2 588.1351, found 582.1357 -7-Methoxy-8-oxo-7-acetamido)-3-((phenylthio)methyl)-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid 21h, Pd2(dba)3·CHCl3 , BIPHEPHOS , and benzenethiol  in MeCN (2 mL) with 10 h of reaction time. Work-up and purification via RP-HPLC  as described above gave the titled compound as a pale-yellow oil . Compound purity by HPLC ; 1H NMR  δ 8.15 , 7.86 , 7.78 , 7.48–7.27 , 5.26 , 4.95 , 4.26 , 3.89 , 3.53 , 3.51 , 3.36 ; 13C NMR  δ 167.0 , 162.5 (C2′), 161.0 (C8), 147.8 , 136.4 (C3), 134.6 (C5″), 132.8 (C6″), 131.4 (C1″), 129.7 (C7″), 129.5 (C3″), 128.7 (C4″), 128.2 (C8″), 126.1 (C2″), 125.2 (C2), 122.9 , 96.7 (C7), 65.6 (C6), 53.8 (O-CH3), 52.5 (CH2-triazole), 37.2 (C3′), 29.1 (C4); IR (cm−1) max 3531 (w), 3280 (w), 3031 (w), 2943 (w), 2840 (w), 1772 , 1704 (s), 1623 (w), 1533 (m), 1482 (w), 1468 (m), 1415 (m), 1371 (m), 1152 (w), 1109 (m), 1087 (m), 1024 (w), 1000 (2), 832 (s), 746 (s), 629 (s); MS (ESI + ve) m/z 538 , 560 , 576 ([M + K]+ 37%); (ESI − ve) m/z 536 , 650 ; HRMS (ESI + ve TOF) calcd for C25H23N5O5S2Na 560.1117, found 560.1122 -3-[(acetyloxy)methyl]-7-amino-7-methoxy-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid. The 7-acetamino moiety was introduced through azidation of the novel α-bromo acetamide 18 followed by a Cu(I)-catalysed azide–alkyne cycloaddition reaction with phenylacetylene, while the 3′-arylthiol substituent was introduced via a palladium-catalyzed arylthioallylation reaction. The chemistry described, and several of the synthetic intermediates reported here, are potentially valuable methods and scaffolds, respectively, for further development of β-lactam antibiotics.The synthesis of eight 7α-methoxy-7-1


Tricholoma ustaloides Romagn., a species belonging to the large Tricholoma genus of higher mushrooms . They were isolated through multiple chromatographic separations, and the structures, including the absolute configuration, were established through a detailed analysis of MS, NMR, and CD spectral data and comparison with related compounds reported in the literature, which has been thoroughly revised.Four novel seconeodolastane diterpenoids, named tricholomalides D–G, were isolated, together with the known tricholomalide C, from the fruiting bodies of  Tricholoma (Fr.) Staude + (C20H28O5)+ (6), 333 [M-CH3]+ (46), 263 (100), 175 (21), 167 (59), 139 (23), 109 (25), 97 (39), 83 (21), 69 (50), 55 (36), 41 (60). Anal. Calcd for C20H28O5: C, 68.94; H, 8.10. Found: C, 69.15; H, 8.18.Colorless, sticky oil; [α]D22 + 31.2 ; Rf 0.53 ; CD (MeOH) Δε220 −0.45, Δε246 +0.34, Δε350 +0.11; 1H and 13C NMR spectral data, see max 2968, 1783, 1736, 1699, 1648, 1367, 1241, 1027, 990, 961 cm−1; EIMS m/z (relative intensity) 388 [M]+ (C22H28O6)+ (54), 359 [M-CHO]+ (11), 346 [M-CH2CO]+ (18), 328 [M-AcOH]+ (34), 313 (12), 285 (45), 269 (21), 263 (33), 178 (23), 163 (48), 133 (37), 106 (78), 95 (33), 83 (42), 69 (35), 58 (69), 43 (100). Anal. Calcd for C22H28O6: C, 66.02; H, 7.27. Found: C, 66.15; H, 7.44.Colorless, sticky oil; [α]D22 + 1.2 ; Rf 0.67 ; CD (MeOH) Δε225 −2.28, Δε246 +0.11, Δε272 −0.68, Δε350 +0.46; 1H and 13C NMR spectral data, see max 3474, 2929, 1780, 1723, 1696, 1647, 1461, 1375, 1246, 1024, 962 cm−1; EIMS m/z (relative intensity) 346 [M]+ (C20H26O5)+ (31), 287 (40), 263 (58), 231 (22), 203 (22), 191 (36), 173 (25), 167 (82), 163 (51), 151 (37), 133 (39), 121 (30), 109 (32), 91 (40), 83 (61), 69 (68), 55 (68), 43 (100). Anal. Calcd for C20H26O5: C, 69.34; H, 7.57. Found: C, 69.75; H, 7.78.Colorless, sticky oil; [α]D22 + 63.9 ; Rf 0.57 ; CD (MeOH) Δε255 +0.47, Δε275 −0.05, Δε340 +0.20; 1H and 13C NMR spectral data, see max 3561, 3416, 2941, 2876, 1783, 1765, 1725, 1638, 1461, 1371, 1242, 1051, 976 cm−1; EIMS m/z (relative intensity) 346 [M]+ (C20H26O5)+ (30), 331 [M-CH3]+ (9), 317 [M-CHO]+ (33), 303 (35), 287 (23), 263 (99), 235 (46), 203 (29), 191 (61), 175 (27), 163 (61), 149 (43), 135 (48), 121 (33), 109 (38), 97 (47), 83 (58), 69 (84), 55 (88), 43 (70), 41 (100). Anal. Calcd for C20H26O5: C, 69.34; H, 7.57. Found: C, 69.65; H, 7.73.Colorless, sticky oil; [α]D22 + 55.2 ; Rf 0.57 ; CD (MeOH) Δε250 +0.43, Δε345 +0.18; 1H and 13C NMR spectral data, see max 3449, 3060, 2960, 2929, 1772, 1731, 1644, 1463, 1370, 1241, 1020, 963, 938, 735 cm−1; EIMS m/z (relative intensity) 390 [M]+ (C22H30O6)+ (9), 375 [M-CH3]+ (54), 315 [M-AcOH]+ (15), 263 (45), 191 (17), 181 (24), 167 (32), 135 (15), 109 (16), 95 (24), 69 (39), 55 (27), 43 (100), 41 (44). Anal. Calcd for C22H30O6: C, 67.67; H, 7.74. Found: C, 67.81; H, 7.62.Colorless, sticky oil; [α]25)  in CH2Cl2 (1 mL) was added to pyridine (50 μL), followed by excess Ac2O (50 μL). The mixture was stirred for 2 h at 22 °C, quenched with MeOH (0.5 mL), and evaporated. The residue was filtered through a short pad of silica gel (300 mg). Elution with CH2Cl2-MeOH at 99:1 gave a product  identical  to tricholomalide D (24).Tricholomalide E (27)  dissolved in CH2Cl2 (0.5 mL). The mixture was stirred for 6 h at 22 °C; subsequently, it was diluted with abundant Et2O. The organic layer was washed with aqueous NaHCO3, dried (Na2SO4), and evaporated. The residue was filtered through a short pad of silica gel (300 mg). Elution with CH2Cl2-MeOH at 99:1 gave a product  identical  to tricholomalide D (24).Freshly prepared PDC   was adde17) was converted to tricholomalide E (25) in 75% yield using the procedure described above.Isolated tricholomalide C .The isolation of four novel seconeodolastane diterpenoids, tricholomalides D–G, from  sordida . InteresTricholoma species grown in the Italian woods, and, regarding tricholomalides C–G, we intend to study their biological activities using in vitro tests.In the future, we will extend our investigations to the chemical contents of other


Simple treatments of partial purification of the reaction crudes  or isolation of the surfactant compositions result in sugar-based compounds having performance levels appropriate to applications in detergency. In addition, the challenging extension of this cascading one-pot synthesis technology to crude milled brown seaweeds was successfully carried out to provide promising surface-active compositions made up of alkyl uronate and alkyl glycoside monosaccharides.The present article describes a one-pot and cascade mode process using biocompatible/biodegradable reagents, for simply obtaining surfactant compositions comprising mixtures of  Currently, 100% biobased surfactants containing both a hydrophobic tail and a hydrophilic head of plant origin represent 5% of the total surfactants in the market worldwide [Fully 100% biobased surfactants on the market mainly include products derived from sugars, which belong to the non-ionic family. They incorporate a glucose head for alkyl polyglucosides (APGs), sucrose for sucro-esters, and sorbitan for sorbitan esters ,3,4. Depd-mannuronate (M) and its C-5 epimer l-guluronate (G). These M and G units are arranged irregularly, by homopolymeric blocks (MM or GG) separated by heterogenous blocks of both uronates (MG) along the alginate chain. Some studies have already shown the possibility of exploiting d-mannuronate oligomers in the surfactant field. However, the preparation of compositions based on monomeric mixtures of both l-guluronate and d-mannuronate units, allowing the valorization of all the sugars present in the biopolymer structure, has not been developed to date.The use of starting materials based on algal polysaccharides, which are characterized by an original chemical functionality compared with polysaccharides from terrestrial plants, constitutes an approach that could make it possible to broaden the fields of application of the 100% biobased surfactants. Alginates, which are polysaccharides present in the cell wall of brown algae, are produced on a scale of 30,000 tons worldwide; their use has several advantages from the point of view of environmental impact, since their production does not require agricultural land, fresh water, or fertilizers and phytosanitary products . These pd-mannopyranosiduronate surfactants have been produced from d-mannuronic acid oligomers through one-pot acid glycosidic bond hydrolysis, esterification, and stereocontrolled Fisher glycosylation in butanol followed by transesterification/transglycosylation processes in fatty alcohols [Alkyl alcohols . These nl-guluronic acid and d-mannuronic acid), and hexoses and pentoses derived from other polysaccharides present in the algal extract . In addition, our objective was to propose an eco-friendly process for the direct conversion of raw algae into n-alkyl glycosiduronic acids and n-alkyl glycosides without the use of organic solvents, i.e., by simultaneously carrying out the steps of extraction, depolymerization by acid hydrolysis with a minimum of water, and esterification/glycosylation. The carboxylic acid-based surfactant compositions produced will be evaluated in terms of surface activities, biodegradability, and ecotoxicity.Within this context, we investigated a novel strategy to synthesize surfactant compositions directly from less refined starting materials to reduce costs and improve the properties expected in the surfactants field . The firLaminaria digitata using a process based on a pre-extraction of the alginates, followed by several steps of precipitation by modulating the pH of the reaction medium in order to separate the G blocks and the M blocks constituting the alginate [Poly(oligo)mannuronates and poly(oligo)guluronates are attractive raw materials for the development of original 100% biobased surfactants. They are obtained from fresh or dry algae derived from alginate ,17. FinaLaminaria digitata, followed by dissolution of the sodium alginates by increasing the pH of the medium. Then, a solid/liquid separation is performed in order to remove the algal residues. At this stage, the liquid fraction can be freeze-dried and constitutes the semi-refined alginates  and semi-refined alginate (s-r Alg) , the prolginates  in the f DP 12.7 .Ascophyllum nodosum species  was used.Regarding the dried milled brown seaweeds, Asco T10 from the In order to achieve the goal of synthesizing novel biobased uronic surfactants from alginates, several uronate sources have been investigated to assess their potential as biomass feedstock for the preparation of such amphiphilic molecules through green synthetic pathways. At first, oligomannuronate (OM) and oligoguluronate (OG) were used in order to define the optimal reaction conditions for the one-pot cascade synthesis of uronate monomeric surfactants. Then the optimized conditions were transposed to oligoalginate with a M/G ratio of 1.4 and a DP of 12.7.1H NMR studies  n-butyl α-d-mannopyranosiduronate 1α was observed as the major monosaccharide product  n-butyl α-d-mannofuranosiduronate 2α (7 mol%), n-butyl α-d-mannofuranosidurono-6,3-lactone 3α (11 mol%), and n-butyl β-d-mannofuranosidurono-6,3-lactone 3β (13 mol%). Products from the guluronate units present in smaller quantities in the raw material (M/G ratio = 2.7) were also identified, i.e., n-butyl-α-l-gulofuranosidurono-6.3-lactone 4α (5 mol%), n-butyl-β-l-gulofuranosidurono-6.3-lactone 4β (5 mol%), and (n-butyl) n-butyl-β-l-gulofuranosiduronate 5β (3 mol%). Alongside these uronate derivatives, a side product 6 was isolated, butyl-5-(dibutoxymethyl)-2-furoate (BDMF) (5 mol%), and it was characterized by NMR  was investigated towards acid hydrolysis, esterification, and Fisher glycosidation conditions. Acid treatment of OM was performed for 7 h with various amounts of methane sulfonic acid (MSA) in the presence of water and butanol . Based o studies . Indeed,d by NMR . This fuuronates . The ove2O (1.7–2 mL/g of crude OM), 140 °C, 7 h) were reproduced and the reaction mixture containing the butyl ester glycoside products 1–5 was subjected directly to simultaneous transesterification and transglycosylation with dodecanol (DodOH). Then the replacement of the butyl chains by the C12 chains was performed in situ  n-dodecyl α-d-mannopyranosiduronate 7α was isolated in addition to several fractions containing mixtures of isomers corresponding to (n-dodecyl) n-dodecyl α-d-mannofuranosiduronate 8α,n-dodecyl α,β-d-mannofuranosidurono-6,3-lactone 9α,β, n-dodecyl-α,β-l-gulofuranosidurono-6.3-lactone 10α,β, (n-dodecyl) n-dodecyl-α,β-l-gulopyranosiduronate 11α,β, and (n-dodecyl) n-dodecyl-β-l-gulofuranosiduronate 12β , H in situ  under re in situ . After anate 12β   that provided dodecyl mannuronate 7α with an overall yield of 25% were thus selected. The molar composition of the mannuronate and guluronate mixture was determined by integrating the characteristic 1H NMR signals of each compound of the uronate mixture (n-dodecyl) n-dodecyl α-d-mannopyranosiduronate 7α (48 mol%), (n-dodecyl) n-dodecyl α-d-mannofuranosiduronate 8α (9 mol%), n-dodecyl α-d-mannofuranosidurono-6,3-lactone 9α (5 mol%), n-dodecyl β-d-mannofuranosidurono-6,3-lactone 9β (22 mol%), n-dodecyl-α-l-gulofuranosidurono-6.3-lactone 10α (3 mol%), n-dodecyl-β-l-gulofuranosidurono-6.3-lactone 10β (3 mol%), (n-dodecyl) n-dodecyl-β-l-gulopyranosiduronate 11β (5 mol%), and (n-dodecyl) n-dodecyl-β-l-gulofuranosiduronate 12β (6 mol%). It is noteworthy that (n-dodecyl) n-dodecyl-α-l-gulopyranosiduronate 11α was not detectable in this mixture due to its presence in quantities that were too small.The comparison of entries 1 and 2  indicate mixture  resultin mixture : , n-dodecyl α-d-mannofuranosidurono-6,3-lactone 9α (4 mol%), n-dodecyl β-d-mannofuranosidurono-6,3-lactone 9β (12% molar), n-dodecyl β-l-gulopyranosiduronic acid 14β (4 mol%), n-dodecyl α-l-gulofuranosidurono-6,3-lactone 10α (9 mol%), n-dodecyl β-l-gulofuranosidurono-6,3-lactone 10β (9 mol%), and n-dodecyl-β-l-gulofuranosiduronic acid 15β (4 mol%), in addition to a non-identified product X (~10 mol%) which could correspond to n-dodecyl d-mannofuranosiduronic acid , n-dodecyl β-l-gulopyranosiduronic acid 14β (13 mol%), n-dodecyl α-l-gulofuranosidurono-6,3-lactone 10α (26 mol%), n-dodecyl β-l-gulofuranosidurono-6,3-lactone 10β (26 mol%), and n-dodecyl-β-l-gulofuranosiduronic acid 15β (21 mol%). The presence of n-dodecyl α,β-d-mannofuranosidurono-6,3-lactone 9α,β was also observed in trace amounts but its quantification was not possible, unlike in the case of synthesis from oligoalginate  as well. It is noteworthy that more MSA (2.2 eq) was required  for OG, nate See . 1H NMR d-mannuronate and l-guluronate units, the same synthetic pathway has been tested with oligoalginate (OAlg) composed of d-Man and l-Gul units with the same reaction conditions used for oligoguluronate (OG) (H–12 OAlgC surfactant composition is characterised by the presence of n-dodecyl α-d-mannopyranosiduronic acid 13α (23 mol%), n-dodecyl α-d-mannofuranosidurono-6,3-lactone 9α (4 mol%), n-dodecyl β-d-mannofuranosidurono-6,3-lactone 9β (9 mol%), n-dodecyl β-l-gulopyranosiduronic acid 14β (9 mol%), n-dodecyl α-l-gulofuranosidurono-6,3-lactone 10α (20 mol%), n-dodecyl β-l-gulofuranosidurono-6,3-lactone 10β (20 mol%), and n-dodecyl-β-l-gulofuranosiduronic acid 15β (9 mol%) (X (6 mol%) formed during the one-pot synthesis of the H–12 ManC composition was also observed. 1H NMR analysis led to the identification of characteristic signals for each compound, as shown in H–12 OAlgC composition are presented in As the process has been shown to be applicable to both ate (OG) . The OAlate (OG) . The fin(9 mol%) . The pres-r Alg) extracted from the Laminaria digitata species composed of d-mannuronate and l-guluronate units in addition to neutral l-fucose, d-glucose, and d-xylose sugars. The first step of the synthetic scheme developed for oligosaccharides could not be directly applied. Indeed, modifications had to be made to take into account the low solubility of the mixture in butanol due to the high degree of polymerization of the polysaccharides. Thus, the synthesis of the butyl C4–C4 derivatives was divided into two steps. First, 2 g of semi-refined alginate containing 810 mg of saccharidic materials was dispersed in water (60 mL) under reflux in the presence of acid  to reduce the degree of polymerization. After 8 h reflux, butanol (60 mL) was added, and the Dean-Stark set-up was used to allow the removal of water by azeotropic distillation (15 h). The overall reaction time (23 h) and the quantity of MSA (5 eq) were increased to optimize the conversion rate  to be protonated without the risk of also protonating the sodium methane sulfonate (pKa = −1.92). It is also to avoid finding methane sulfonic acid in the filtrate that the pH of the aqueous solution should not be decreased below 2. This cascading one-pot method with a final purification step is applicable on an industrial scale and it allows the elimination of fatty alcohols in addition to salts. 1H NMR analysis of the H–12 s-r AlgC composition revealed the presence of the following products (n-dodecyl α-d-mannopyranosiduronic acid 13α (23 mol%), n-dodecyl α-d-mannofuranosidurono-6,3-lactone 9α (5 mol%), n-dodecyl β-d-mannofuranosidurono-6,3-lactone 9β (13 mol%), n-dodecyl β-l-gulopyranosiduronic acid 14β (11 mol%), n-dodecyl β-l-gulofuranosidurono-6,3-lactone 10β (20 mol%), n-dodecyl-α-l-fucopyranosides 17α (5 mol%), n-dodecyl-β-l-fucopyranosides 17β (8 mol%), and n-dodecyl-α,β-l-fucofuranosides 18α,β (15 mol%)  . 1H NMR Ascophyllum nodosum species, Thorverk, Iceland). Indeed, the use of such crude seaweeds could contribute to lowering the production cost of surfactant compositions due to the reduced price of these raw materials compared to refined or semi-refined polysaccharides and to a simplification of the process which no longer requires the prior extraction of alginates.The final challenge was to extend the cascading one-pot synthesis technology developed from oligo- and polysaccharide alginates to commercial milled brown seaweeds rich in alginate and fucoidan polysaccharides  at a temperature of around 90 °C. Using this technology, octanol was recovered in a very efficient way while minimizing the risk of degradation of the sugar derivatives present in the final mixture.The second step of the process based on transesterification/transglycosylation reactions led to the replacement of butyl chains by longer chains using a fatty alcohol as well as the recycling of butanol through a vacuum distillation. Octanol was selected as the fatty alcohol to obtain C1H NMR analysis in CD3OD was also performed, based on the integration of the anomeric sugar proton signals in the 4.2–5.0 ppm region , but it lowers the surface tension much more effectively at low concentration values. This indicates, therefore, that the presence of H–12 ManC isomers allows for improved surfactant performance. The mixture from oligoguluronate gives the highest CMC with a value of 0.12 g L−1 and a γCMC of 28.8 mN m−1. However, for concentrations below 0.008 g L−1, the lowest surface tension values were measured with this H–12 GulC composition. Similar CMC and γCMC values were obtained with the batch from oligoalginate . Finally, the surfactant mixture obtained from semi-refined alginate provided the lowest CMC with a value of 0.04 g L−1 and a γCMC of 29.0 mN m−1. Tensiometry measurements were also carried out on anionic surfactant, sodium laureth sulfate (SLES). The CMC was determined at 0.13 g L−1 and the γCMC at 31.9 mN m−1. Thus, all the compounds obtained from the different sources of algae extracts exhibit better performance in terms of CMC and γCMC.The different batches of H-Curements . The resacid 13α  exhibitsH–8 crude seaweedC) revealed a CMC value of 0.007 g L−1 and a γCMC of 27.0 mN m−1, demonstrating excellent surfactant properties.Surprisingly, surface tension measurements in pH 7 buffered solution from the composition derived from crude seaweed and octanol (H–12 s-r AlgC) in addition to standard SLES. Experiments to determine the ecotoxicity of products include effects on aquatic organisms, from microalgae to fish. These different pollutant-sensitive organisms serve as controls. Three standardized tests were conducted: an algal growth inhibition test, a microcrustaceous immobilization test, and a lethal toxicity test on freshwater fish. The microalgae used, called Pseudokirchneriella subcapitata, are ubiquitous in the environment and are sensitive to toxic substances. The test is conducted according to the OECD 201 method [Brachydanio rerio test population after a 96-h exposure period (CL50). Brachydanio rerio is one of the model organisms commonly encountered in research laboratories for fish behavior studies [A series of ecotoxicity studies have been performed with the uronic surfactant compositions derived from semi-refined alginate and dodecanol . The CO2 formed during degradation is trapped in external containers. The tests are conducted over 28 days, during which the evolution of the biodegradation rate is determined. The OECD 301 B method considers a product to be readily biodegradable if the biodegradation rate has reached at least 60% after 28 days and readily biodegradable with respect to the 10-day period if the biodegradation rate has reached at least 60% rate 10 days after the rate has reached 10%.The comparative aerobic readily biodegradability of compounds B method . The objH–12 s-r AlgC, exhibited lower biodegradability rates after 28 days (72%) than SLES (94%). Under the experimental conditions of the test, H–12 s-r AlgC, . In contrast, for SLES, the CO2 release threshold of 60% of the theoretical CO2 was reached within 10 days , and filtration steps allows the final compositions to be enriched with active ingredients by removing  residual salts and fatty alcohols. In terms of performance, the carboxylic acid-based surfactant compositions allow surface tension to be lowered to values ≤30 mN m−1 at concentrations equivalent to or lower than that of sodium laureth sulfate. In particular, the compositions derived from semi-refined alginate (s-r Alg) or oligomannuronate (OM) lead to CMC values two to three times lower than that obtained from sodium laureth sulfate. In addition, a significant improvement in ecotoxicity profile was observed for the biodegradable surfactant composition H–12 s-r AlgC, especially towards Daphnia.To facilitate the industrial development of innovative surfactants based on carboxylate saccharides, we have proposed a new approach to produce these surfactants without sulfate and without ethylene oxide, according to a smart strategy based on a multi-step process carried out in a cascading single-pot mode without separating and isolating any reaction intermediates and applicable to oligomeric or polymeric alginates in a more or less refined form. The innovation of this research is also based on its positioning in green/blue chemistry . The syntheses involve a succession of chemical reactions such as depolymerization by acid hydrolysis of oligo- or polysaccharides, esterification and glycosylation with H–8 crude AlgC is rich in carboxylic acid sugars, which also contain alkyl glycosides derived from neutral sugars representative of the structure of the polysaccharides present in the starting plant material (d-fucose). These modify the physicochemical properties of the final compositions and optimise their surface-active performance [−1) and critical micellar concentration (0.007 g L−1) were obtained in the case of these mixtures of uronic acid- and fucose-based surfactants.In order to further simplify the process and reduce costs, a novel strategy has been successfully tested based on the direct use of plant materials in their raw state, i.e., without any chemical transformation, in particular solvent extraction or enzymatic extraction, and at most a mechanical and/or physical transformation, such as washing, grinding, and/or drying. A notable advantage of this approach is the use of plant materials with a water content of up to 13% by weight, thus avoiding the addition of water during the depolymerization step by hydrolysis of the polysaccharides. The process has been applied to brown seaweed containing alginates and fucans and/or fucoidans. The process incorporates the same reaction sequences as for polysaccharides  with the addition of a preliminary extraction step carried out in situ as well as filtration, centrifugation, or distillation steps under reduced pressure, allowing the elimination of the residues of plant material not involved in the syntheses and the recycling of the excess alcohols. The obtained composition formance . Indeed,254 Merck). Compounds were visualized using a H2SO4 solution (5% H2SO4 in EtOH) or a vanillin solution (15 g of vanillin in 250 mL of EtOH and 2.5 mL of conc. H2SO4) followed by heating. Geduran 60  was used for column chromatography. NMR spectra were recorded on a Bruker Avance III 400 spectrometer operating at 400.13 MHz for 1H, equipped with a BBFO probe with a Z-gradient coil and a GREAT 1/10 gradient unit. The standard temperature was adjusted to 298 K. The zg30 Bruker pulse program was used for 1D 1H NMR, with a TD of 64k, a relaxation delay d1 = 2 s, and 8 scans. The spectrum width was set to 18 ppm. Fourier transform of the acquired FID was performed with an apodization of 0.3 Hz in most of the cases. Chemical shifts are mentioned in parts per million (ppm) with tetramethylsilane as an internal standard. Coupling constants were expressed in Hertz (Hz) and the following abbreviations were used to indicate the multiplicity: s (singulet), d (doublet), t (triplet), q (quadruplet), m (multiplet), dd (doublet of doublets), dt (doublet of triplets), and br .Oligomannnuronate, oligoguluronate, oligoalginate, and semi-refined alginate were produced from fresh or dry seaweeds  ,17,18. D4C–4 ManC from OMPreparation of −2, 1 eq.) was dispersed in water (0.87 mL) and n-butanol (25 mL) in a round-bottom flask with a Dean-Stark apparatus. Technical-grade methanesulfonic acid 70% wt  was added and the mixture was refluxed under vigorous stirring. The water formed in the medium was gradually removed by azeotropic distillation. After 7 h, the mixture was cooled to ambient temperature. The mixture was then neutralized with 1N NaOH solution (400 µL) and concentrated. The resulting mixture was dissolved in diethyl ether (20 mL) and then filtered using celite. The celite was finally rinsed with diethyl ether (100 mL) and the filtrate was concentrated. The products obtained (495 mg) were purified by silica gel chromatography with CH2Cl2:CH3OH,  to help determine the molar composition. The molar composition of this mixture of products before silica gel chromatography is as follows: 52% (n-butyl) n-butyl-α-d-mannopyranosiduronate (1α), 7% (n-butyl) n-butyl-α-d-mannofuranosiduronate (2α), 11% n-butyl-α-d-mannofuranosidurono-6.3-lactone (3α), 13% n-butyl-β-d-mannofuranosidurono-6.3-lactone (3β), 5% BDMF (6), 5% n-butyl-α-l-gulofuranosidurono-6.3-lactone (4α), 5% n-butyl-β-l-gulofuranosidurono-6.3-lactone (4β), 3% (n-butyl) n-butyl-β-l-gulofuranosiduronate (5β). 1H NMR  δ 7.11 , 6.51 , 5.53 , 5.14 , 5.06 , 5.05 , 5.03 , 5.00 , 4.95 , 4.91 , 4.90 , 4.79 , 4.76 , 4.72 , 4.69 , 4.43 ,4.31–4.15 , 4.08 , 4.00 , 3.94 , 3.87 , 3.83–3.69 , 3.65 , 3.60–3.38  1.76–1.62 , 1.62–1.49 , 1.46–1.29 , 0.99–0.88 . After column chromatography, five fractions F1–F5 were isolated and characterised by 1H NMR. F1: 57 mg of BDMF (6). ): 1H NMR  δ 7.11 , 6.51 , 5.53 (s), 4.36 , 3.65 , 1.80–1.63 , 1.63–1.50 , 1.48–1.31 , 1.00–0.86 ; F2: 12 mg of n-butyl-α-l-gulofuranosidurono-6.3-lactone (4α) and (n-butyl) n-butyl-β-l-gulofuranosiduronate (5β). ): 1H NMR  δ 4.98 , 4.95 , 4.91 , 4.69 , 4.52 , 4.38 , 4.28 , 4.26–4.18 , 4.15 , 3.91 , 3.71 , 3.58 , 3.43 , 1.73–1.59 , 1.58–1.47 , 1.46–1.24 , 0.99–0.86 ; F3: 65 mg of n-butyl-β-d-mannofuranosidurono-6.3-lactone (3β), (n-butyl) n-butyl-α-d-mannofuranosiduronate (2α), n-butyl-α-l-gulofuranosidurono-6.3-lactone (4α), n-butyl-β-l-gulofuranosidurono-6.3-lactone (4β) and (n-butyl) n-butyl-β-l-gulofuranosiduronate (5β). ): 1H NMR  δ 5.14 , 5.06 , 5.03 , 4.97 , 4.95 , 4.90 , 4.79 , 4.72 , 4.69 , 4.56 , 4.52 , 4.43 , 4.39 , 4.30–4.19 , 3.93 , 3.79 , 3.66 , 3.54–3.38 , 1.73–1.47 , 1.45–1.24 , 0.93 ; F4: 49 mg of n-butyl-β-d-mannofuranosidurono-6.3-lactone (3β), n-butyl-α-d-mannofuranosidurono-6.3-lactone (3α) and n-butyl-β-l-gulofuranosidurono-6.3-lactone(4β). ): 1H NMR  δ 5.13 , 5.05 , 5.03 , 5.00 , 4.90 , 4.79 , 4.76 , 4.72 , 4.27–4.17 , 3.74–3.61 , 3.50–3.35 , 1.70–1.46 , 1.42–1.27 , 0.96–0.86 ; F5: 183 mg of (n-butyl) n-butyl-α-d-mannopyranosiduronate (1α). ): 1H NMR  δ 4.90 , 4.28–4.15 , 4.28–4.15 , 4.08 , 4.00 , 3.93 , 3.86 , 3.72 , 3.50–3.43 , 1.72–1.62 , 1.62–1.51 , 1.44–1.30 , 0.93 .Oligomannuronate  was dispersed in water (0.9 mL) and n-butanol (25 mL) in a round-bottom flask with a Dean-Stark apparatus. Methanesulfonic acid technical grade 70% wt  was added and the mixture was refluxed under vigorous stirring. The water formed in the medium was gradually removed by azeotropic distillation. After 7 h, the mixture was cooled to ambient temperature. Dodecanol  and the 70% methanesulfonic acid solution  were added. The mixture was stirred at 70 °C under reduced pressure (up to 5 mbar) using distillation apparatus. Once the butanol had been completely removed (2.17 h), water (16 mL) was added and the mixture was then neutralized with 1N NaOH solution (0.5 mL). The whole mixture was left to stir vigorously at 80 °C for 1 h and the mixture was cooled to ambient temperature. The mixture was cooled at 0 °C to solidify the organic phase which was isolated by solid-liquid extraction and dried under vacuum. The resulting mixture (1.684 g) was purified by a first silica gel chromatography with CH2Cl2/CH3OH, (100:0–96:4) to give crude mixture 12C–12 ManC (504 mg). The molar composition of this mixture is as follows: 48% (n-dodecyl) n-dodecyl-α-d-mannopyranosiduronate (7α), 9% (n-dodecyl) n-dodecyl-α-d-mannofuranosiduronate (8α), 5% n-dodecyl-α-d-mannofuranosidurono-6.3-lactone (9α), 22% n-dodecyl-β-d-mannofuranosidurono-6.3-lactone (9β), 3% n-dodecyl-α-l-gulofuranosidurono-6.3-lactone (10α), 3% n-dodecyl-β-l-gulofuranosidurono-6.3-lactone (10β), 5% (n-dodecyl) n-dodecyl-β-l-gulopyranosiduronate (11β), 6% (n-dodecyl) n-dodecyl-β-l-gulofuranosiduronate (12β). 1H NMR  δ 5.07 , 5.05 , 5.03 , 4.99 , 4.97 , 4.97 , 4.95 , 4.92 , 4.91 , 4.90 , 4.79 , 4.72 , 4.69 , 4.58 , 4.55 , 4.53 , 4.43 , 4.39 , 4.29–4.14 , 4.09 , 4.00 , 3.95 , 3.89 , 3.82–3.68 , 3.53–3.37 , 1.74–1.64 , 1.63–1.53 , 1.38–1.20 , 0.91–0.85 . A second silica gel chromatography with CH2Cl2/CH3OH, (97:3) was made to help determine the molar composition. Five fractions F1–F5 were isolated and characterised by 1H NMR. F1: 74 mg of (n-dodecyl) n-dodecyl-α-d-mannofuranosiduronate (8α), n-dodecyl-β-d-mannofuranosidurono-6.3-lactone (9β), n-dodecyl-α-l-gulofuranosidurono-6.3-lactone (10α) and (n-dodecyl) n-dodecyl-β-l-gulopyranosiduronate (12β). ): 1H NMR  δ 5.03 , 4.98 , 4.97 , 4.96 , 4.91 , 4.89 , 4.79 , 4.70 , 4.71–4.65 , 4.65–4.61 , 4.52 , 4.43 , 4.37 , 4.27–4.18 , 3.92 , 3.81–3.54 , 1.76–1.63 , 1.62–1.48 , 1.39–1.20 , 0.91–0.85 ; F2: 64 mg of n-dodecyl-α-d-mannofuranosiduronate (8α), n-dodecyl-β-d-mannofuranosidurono-6.3-lactone (9β). ): 1H NMR  δ 5.03 , 4.97 , 4.90 , 4.79 , 4.62 , 4.42 , 4.38 , 4.29–4.18 , 3.92 , 3.78 , 3.65 , 3.53–3.35 ), 1.74–1.63 , 1.63–1.50 , 1.38–1.19 , 0.91–0.85 ; F3: 30 mg of (n-dodecyl) n-dodecyl-α-d-mannopyranosiduronate (7α), (n-dodecyl) n-dodecyl-α-d-mannofuranosiduronate (8α), n-dodecyl-α-d-mannofuranosidurono-6.3-lactone (9α), n-dodecyl-β-d-mannofuranosidurono-6.3-lactone (9β), n-dodecyl-β-l-gulofuranosidurono-6.3-lactone (10β), (n-dodecyl) n-dodecyl-α-l-gulopyranosiduronate (11α), (n-dodecyl) n-dodecyl-β-l-gulopyranosiduronate (11β), and (n-dodecyl) n-dodecyl-β-l-gulofuranosiduronate (12β). ): 1H NMR  δ 5.14 , 5.07 , 5.05 , 5.03 , 4.99 , 4.97 , 4.92 , 4.91 , 4.89 , 4.78 , 4.72 , 4.65–4.61 , 4.63 , 4.60 , 4.57 , 4.55 , 4.52 , 4.43 , 4.40 , 4.39 , 4.27–4.18 , 4.13 , 4.09 , 3.80–3.64 , 3.55–3.37 , 1.73–1.48 , 1.42–1.18 , 0.93–0.84 ; F4: 240 mg of (n-dodecyl) n-dodecyl-α-d-mannopyranosiduronate (7α). ): 1H NMR  δ 4.91 , 4.21 , 4.08 , 4.01 , 3.94 , 3.88 , 3.72 , 3.47 , 1.73–1.64 , 1.64–1.52 , 1.38–1.22 , 0.92–0.84 ; F5: 9 mg of a mixture of products .Oligomannuronate  was dispersed in water (1 mL) and n-butanol (25 mL) in a round-bottom flask with a Dean-Stark apparatus. Methanesulfonic acid technical grade 70% wt  was added and the mixture was refluxed under vigorous stirring. The water formed in the medium was gradually removed by azeotropic distillation. After 7 h, the mixture was cooled to ambient temperature. Dodecanol  and the 70% methanesulfonic acid solution  were added. The mixture was stirred at 70 °C under reduced pressure (up to 5 mbar) using distillation apparatus. Once the butanol had been completely removed (1.42 h), a 0.4N NaOH solution  was added and the mixture was left to stir vigorously at 70 °C. for 1 h. The water was then removed by freeze-drying or by azeotropic distillation with butanol. The excess dodecanol present in the crude product was removed by solid-liquid extraction with EtOAc. At the end of this treatment, the mixture of products was dissolved in ice-cold water (15 mL) and EtOAc (22.5 mL) followed by a 1N hydrochloric acid solution (3.1 mL) was added. The aqueous solution was extracted with EtOAc (5 × 7.5 mL). The organic phases were combined and washed with a saturated NaC1 solution (30 mL) and a 1N hydrochloric acid solution (150 µL). The organic phase was dried with MgSO4 and then concentrated under vacuum. A mixture of products H–12 ManC was obtained (341 mg), the molar composition of which is: 47% n-dodecyl-α-d-mannopyranosiduronic (13α), 4% n-dodecyl-α-d-mannofuranosidurono-6.3-lactone (9α), 12% n-dodecyl-β-d-mannofuranosidurono-6.3-lactone (9β), 4% n-dodecyl-β-l-gulopyranosiduronic (14β), 9% n-dodecyl-α-l-gulofuranosidurono-6,3-lactone (10α), 9% n-dodecyl-β-l-gulofuranosidurono-6,3-lactone (10β), 4% n-dodecyl-β-l-gulofuranosiduronic acid (15β), 10% non-identified molecule X. 1H NMR  δ 5.11 , 5.07 , 5.00 , 4.98 , 4.97 , 4.96 , 4.92 , 4.80 , 4.73 , 4.62 , 4.62–4.56 , 4.55 , 4.49 , 4.47 , 4.43–4.38 (m), 4.35 , 4.29 , 4.18 , 4.14–4.08 , 4.05 , 4.00 , 3.89 , 3.79 , 3.76–3.63 , 3.56–3.38 , 1.67–1.51 , 1.46–1.22 , 0.94–0.87 .Oligomannuronate  was dispersed in water (1.5 mL) and n-butanol (29 mL) in a round-bottom flask with a Dean-Stark apparatus. Methanesulfonic acid technical grade 70% wt  was added and the mixture was refluxed under vigorous stirring. The water formed in the medium was gradually removed by azeotropic distillation. After 7 h, the mixture was cooled to ambient temperature. Dodecanol  and the 70% methanesulfonic acid solution  were added. The mixture was stirred at 70 °C under reduced pressure (up to 5 mbar) using distillation apparatus. Once the butanol had been completely removed (1.42 h), a 0.4N NaOH solution  was added and the mixture was left to stir vigorously at 70 °C. for 1 h. The water was then removed by freeze-drying or by azeotropic distillation with butanol. The excess dodecanol present in the crude product was removed by solid-liquid extraction with EtOAc. At the end of this treatment, the mixture of products was dissolved in ice-cold water (27.5 mL) and EtOAc (27.5 mL), then a 1N hydrochloric acid solution (3.6 mL) was added. The aqueous solution was extracted with EtOAc (3 × 15 mL). The organic phases were combined and washed with a saturated NaC1 solution (35 mL). The organic phase was dried with MgSO4 and then concentrated under vacuum. A mixture of products H–12 GulC was obtained (356 mg), the molar composition of which is: 15% n-dodecyl α-d-mannopyranosiduronic acid (13α), 13% n-dodecyl β-l-gulopyranosiduronic acid (14β), 26% n-dodecyl α-l-gulofuranosidurono-6,3-lactone (10α), 26% n-dodecyl β-l-gulofuranosidurono-6,3-lactone (10β), 21% n-dodecyl-β-l-gulofuranosiduronic acid (15β). 1H NMR  δ 5.07 , 4.98 , 4.96 , 4.92 , 4.80 , 4.62 , 4.51 , 4.49 , 4.39 , 4.29 , 4.02 , 4.00 , 3.89 , 3.84 , 3.79 , 3.76–3.59 , 3.56–3.36 , 1.69–1.50 , 1.45–1.25 , 0.97–0.87 .Oligoguluronate  was dispersed in water (2.0 mL) and n-butanol (39 mL) in a round-bottom flask with a Dean-Stark apparatus. Methanesulfonic acid technical grade 70% wt  was added and the mixture was refluxed under vigorous stirring. The water formed in the medium was gradually removed by azeotropic distillation. After 7 h, the mixture was cooled to ambient temperature. Dodecanol  and the 70% methanesulfonic acid solution  were added. The mixture was stirred at 70 °C under reduced pressure (up to 5 mbar) using distillation apparatus. Once the butanol had been completely removed (1.42 h), a 0.4N NaOH solution (25 mL) was added and the mixture was left to stir vigorously at 70 °C. for 1 h. The water was then removed by freeze-drying or by azeotropic distillation with butanol. The excess dodecanol present in the crude product was removed by solid-liquid extraction with EtOAc. At the end of this treatment, the mixture of products was dissolved in ice-cold water (30 mL) and EtOAc (45 mL), then a 1N hydrochloric acid solution (6 mL) was added. The aqueous solution was extracted with EtOAc (8 × 15 mL). The organic phases were combined and washed with a saturated NaC1 solution (60 mL) and a 1N hydrochloric acid solution (300 µL). The organic phase was dried with MgSO4 and then concentrated under vacuum. A mixture of products H–12 OAlgC was obtained (759 mg), the molar composition of which is: 23% n-dodecyl-α-d-mannopyranosiduronic (13α), 9% n-dodecyl-β-l-gulopyranosiduronic (14β), 20% n-dodecyl-α-l-gulofuranosidurono-6,3-lactone (10α), 20% n-dodecyl-β-l-gulofuranosidurono-6,3-lactone (10β), 9% n-dodecyl-β-l-gulofuranosiduronic acid (15β), 4% n-dodecyl-α-d-mannofuranosidurono-6.3-lactone (9α), 9% n-dodecyl-β-d-mannofuranosidurono-6.3-lactone (9β), 6% non-identified molecule X. 1H NMR  δ 5.11 , 5.07 , 4.97 , 4.95 , 4.95 , 4.91 , 4.80 , 4.72 , 4.61 , 4.60–4.55 , 4.54 , 4.51 (s (br), H4-15β), 4.48 , 4.46 , 4.38 , 4.35 , 4.29 , 4.12–4.08 , 3.88 , 3.78 , 3.75–3.61 , 3.55–3.35 , 1.70–1.49 , 1.45–1.23 , 0.95–0.86 .Oligoalginate  was dispersed in water (60 mL) and the 70% methane-sulfonic acid solution  was added in a round-bottom flask with a Dean-Stark apparatus. The mixture was refluxed with vigorous stirring. At 8 h of reaction, butanol (60 mL) was added and the mixture was left at reflux with vigorous stirring. The water present in the medium was gradually removed by azeotropic distillation. After a further 15 h of reaction, and once the mixture had returned to ambient temperature, dodecanol  was added. The mixture was stirred at 70 °C under reduced pressure (up to 5 mbar) using distillation apparatus. Once the butanol had been completely removed (1.42 h), a 0.2 N NaOH solution (60 mL) was added and the mixture was left to stir vigorously at 70 °C. for 1 h. The water was then removed by freeze-drying or by azeotropic distillation with butanol. The excess dodecanol present in the crude product was removed by solid-liquid extraction with acetone. At the end of this treatment, the mixture of products was dissolved in ice-cold water (75 mL) and then a 0.5 M oxalic acid solution (8.0 mL) was added. The water was then removed by freeze-drying. The crude product was purified by solid-liquid extraction with acetone (20 mL + 8 × 10 mL). The filtrate was concentrated under vacuum. At the end of this treatment, a mixture of products was obtained (628 mg), the weight composition of which is: 23% n-dodecyl-α-d-mannopyranosiduronic (13α), 5% n-dodecyl-α-d-mannofuranosidurono-6.3-lactone (9α), 13% n-dodecyl-β-d-mannofuranosidurono-6.3-lactone (9β), 11% n-dodecyl-β-l-gulopyranosiduronic (14β), 20% n-dodecyl-β-l-gulofuranosidurono-6,3-lactone (10β), 5% n-dodecyl-α-l-fucopyranosides 17α, 8% n-dodecyl-β-l-fucopyranosides 17β and 15% n-dodecyl-α,β-l-fucofuranosides 18α,β. 1H NMR  δ 5.08 , 5.03 , 4.95 , 4.89 , 4.85 , 4.83 , 4.80 , 4.75 , 4.65 , 4.56 , 4.51 , 4.43 , 4.27 , 4.25 , 4.18 , 4.09 , 3.83 , 3.79–3.63 , 3.51–3.31 , 1.68–1.49 , 1.46–1.26 , 1.24 , 1.22 , 1.21 , 0.97–0.86 .The semi-refined alginate derived from H–8 crude AlgC from crude AlgPreparation of Ascophyllum nodosum seaweed (100 g) was introduced into the reactor (1 L). Butanol (140 mL) and the methane-sulfonic acid solution (30 g of 70% MSA in 140 mL of butanol) were added. The mixture was refluxed and stirred with a four-blade Teflon paddle at 400 rpm. For the double envelope, the temperature was fixed at 135 °C. After 7 h of reaction, the mixture was recovered hot through the lower tap and then filtered or centrifuged to remove residues. The presence of products in the reaction medium was monitored by TLC with CH2Cl2/MeOH  as the eluent and vanillin solution as the staining reagent. The filtrate/supernatant was transferred into the reactor (1 L) and octanol (100 mL) was added. The reaction mixture is pulled under vacuum at 70 °C for 5–7 h to evaporate butanol and perform the transesterification/transglycosylation reactions. The mixture was made alkaline with a concentrated sodium hydroxide solution . After 1–2 h at 70 °C, only non-ionic surfactants were visible by TLC ). The anionic molecules were revealed by TLC ). The mixture was pulled under vacuum to evaporate the water. At room temperature, 95% of H2SO4 was added to acidify to pH 2–3 and precipitate the salts (counter-ions of the anionic surfactants). Surfactants were miscible in octanol. This oil phase was filtered to remove the salts which were insoluble. The oily filtrate was injected into a molecular distillation apparatus to eliminate octanol. At 100 °C under reduced pressure (1 mbar), 24 g of the surfactant composition was recovered in the form of a viscous syrup, and consequently with a mass yield of approximately 24% relative to the starting algae. At room temperature, the product is a pasty and hygroscopic solid. The mixture was submitted to thermogravimetric analysis, which showed that it consisted of 83.2% organic matter; 10.5% water; and 6.75% ash. 1H NMR analysis revealed the presence of 30 mol% n-octyl α-d-mannopyranosiduronic acid 19α, 6 mol% n-octyl β-l-gulopyranosiduronic acid 20β, n-octyl α-l-gulofuranosidurono-6,3-lactone 21α, 13 mol% n-octyl-α-l-fucopyranoside 22α, 9 mol% n-octyl-β-l-fucopyranosides 22β, 9 mol% n-ocyl-α-l-fucofuranosides 23α and 28 mol% n-octyl-β-l-fucofuranoside 23β. 1H NMR  δ 5.05 , 4.97 , 4.93 , 4.82 , 4.78 , 4.75 , 4.72 , 4.64 , 4.61 , 4.50 , 4.43 , 4.26 , 3.88 , 3.80 , 3.77–3.63 , 3.53–3.35 , 1.72–1.50 , 1.48–1.30 , 1.27 , 1.26 , 1.22 , 1.19 , 1.01–0.88 .The dried milled Critical micelle concentrations (CMC) and interfacial tensions (IFT) were measured on a force tensiometer Krüss K100. The critical micelle concentrations (CMC) were determined using the du Noüy ring method and the Krüss Laboratory Desktop software with the Surfactant Characteristics program. A water solution of surfactant was prepared around 10 g/L. Only 10 mL of this solution was used for the CMC determination. The deionized water was added thanks to an automatic burette controlled by the software. The concentration and tension determination were automatically determined by the software. The CMC was measured at 25 °C.In this study, it has been proven for the first time that the conversion of oligo- and polysaccharide alginates in refined or semi-refined forms into biodegradable alkyl uronate surfactants can be carried out by one-pot acid hydrolysis, butanolysis, transesterification, transacetalisation, and saponification reactions, thus avoiding the isolation of any reaction intermediates. In addition, an in situ process was developed to manufacture surfactant compositions directly from crude milled brown seaweeds without requiring the standard steps of polysaccharide extraction and purification. These alkyl uronate monosaccharides as isomeric mixtures, with or without alkyl glycoside co-products, exhibit attractive surface activity and reduced aquatic ecotoxicity compared to commercial sodium laureth sulphate. Further physicochemical studies to evaluate the potential of these innovative surfactant compositions in cosmetic formulations are currently under investigation. Moreover, the transposition of the strategy, which consists in using crude vegetable raw materials instead of extracted polysaccharides, was also successfully achieved in the case of sugar beet pulps containing pectins ,26: this