Scalable Synthesis and Cancer Cell Cytotoxicity of Rooperol and Analogues.

Plant polyphenols, such as the African potato (Hypoxis hemerocallidea)-derived bis-catechol rooperol, can display promising anticancer activity yet suffer from rapid metabolism. Embarking upon a program to systematically examine potentially more metabolically stable replacements for the catechol rings in rooperol, we report here a general, scalable synthesis of rooperol and analogues that builds on our previous synthetic approach incorporating a key Pd-catalyzed decarboxylative coupling strategy. Using this approach, we have prepared and evaluated the cancer cell cytotoxicity of rooperol and a series of analogues. While none of the analogues examined here were superior to rooperol in preventing the growth of cancer cells, analogues containing phenol or methylenedioxyphenyl replacements for one or both catechol rings were nearly as effective as rooperol.

1. Introduction

African potato (Hypoxis hemerocallidea) is a widely used medicinal plant in southern Africa [1]. Ethanolic extracts of the corms of H. hemerocallidea contain 10–15% of hypoxoside, a bis-glucoside of the aglycone rooperol, 1 (Figure 1) [1]. Rooperol has demonstrated a cytotoxicity against cancer cell lines [2,3,4], and an orally administered extract of H. hemercallidea has been the subject of Phase I clinical trial in advanced lung cancer patients [5,6]. Interestingly, in this Phase I trial, no dose-limiting toxicity was identified, and 5 of the 24 patients enrolled showed some signs of response, including one complete response (>5 years) [6]. Despite this hint of clinical promise, pharmacokinetic studies demonstrated the very rapid metabolism of hypoxoside and rooperol, with only Phase II metabolites of rooperol detected in the blood of these patients [5].

Figure 1

Structure of rooperol (top) and prior synthetic scheme for rooperol analogues 1 (bottom).

Rooperol is one of a number of plant polyphenols that have demonstrated promise as anticancer agents [7,8,9]. However, a common issue with these anticancer polyphenols is metabolic instability [10]. Our work has focused on the design and evaluation of analogues of biologically interesting plant polyphenols with the goal of improved metabolic stability [11,12]. In the present case, our efforts focused on rooperol analogues with more metabolically stable replacements for this natural product’s catechol moieties.

A number of synthetic studies of rooperol and analogues have been reported. The first total synthesis by Drewes and co-workers employed a problematic Csp-Csp3 coupling step [13]. We subsequently reported a very concise synthesis of rooperol and analogues, which incorporated two key strategies: a Friedel–Crafts reaction involving tetrachlorocyclopropene and electron-rich aromatics and a Pd-catalyzed decarboxylative coupling of the resulting 3-arylprop-2-ene-1-yl arylpropiolates 2 (Figure 1) [14]. Although this route could afford rooperol analogues in as few as three steps, it suffers from two limitations. First, the route shown in Figure 1 lacks generality in that only very electron-rich aromatic compounds participate in the initial Friedel–Crafts reaction, and only certain arylprop-2-ene-1-ols directly afford the esters 2 upon alcoholysis of the initially formed Friedel–Crafts product. Second, the Friedel–Crafts reaction did not proceed well above 1 mmole scale, limiting the amount of material that could be prepared for biological assay. Thus, in order to continue our efforts to prepare and evaluate metabolically stable rooperol analogues, we first had to redesign this synthesis. Here, we report a more general, scalable synthesis of rooperol and analogs that retain the key Pd-catalyzed decarboxylative coupling from our earlier work. Using this improved synthesis, we prepared a number of rooperol analogs and report here their activity against HeLa cancer cells.

2. Results

We set out to evaluate a variety of potential replacements for the catechol moieties of rooperol. As shown in Figure 2, these replacements included variations in the number (B) and placement (C) of the catechol hydroxyl groups and substitution of these groups with fluorine (D) or methylenedioxy groups (E). In addition, we also explored the 4H-benzo[d][1,3]dioxine group (F) as a replacement for the catechol groups in rooperol.

Figure 2

Replacements for the catechol groups (A) of rooperol.

Given the limitations of our previous route to rooperol and analogues (Figure 1), the preparation of the analogues contemplated in Figure 2 required an alternative synthetic scheme. Our revised synthesis of protected rooperol and analogs is shown in Scheme 1. We prepared arylprop-2-ene-1-ols 5 from the corresponding aldehydes 8 via reduction of the methyl esters 4, obtained via Wittig reaction. Separately, the same aldehydes 8 were subjected to Corey–Fuchs alkynylation [15] via the vinyl dibromides 6, which were subjected to elimination with nBuLi, followed by trapping of the resulting alkynyl anions with CO2 to afford the arylpropiolic acids 7. All of these transformations occurred without incident to afford good yields of the products 4A–F, 5A–F, 6A–E, and 7A–E with the exception of the nBuLi elimination/trapping of the vinyl dibromide 6D derived from 3,4-difluorobenzaldehyde. This reaction proved somewhat capricious, and at best, only modest yields of the corresponding acid 7D were obtained.

Scheme 1

Scalable and general synthesis of protected rooperol (1′AA) and analogues.

The preparation of the carboxylic acid 7F followed a different route, as shown in Scheme 2. The previously reported alkyne 9F [16] was deprotonated with nBuLi, and the resulting anion was trapped with CO2 to afford 7F in good yield. While the aldehydes 8D and 8E are commercially available, the tert-butyldimethylsilyl-protected aldehydes 8A–C were prepared by reacting the commercially available hydroxy-substituted benzaldehydes 10A–C with tert-butyldimethylsilyl chloride in the presence of imidazole (Scheme 2). The previously reported aldehyde 8F was prepared from the corresponding bromide 11F following literature precedent [16] (Scheme 2).

Scheme 2

Preparation of carboxylic acid 7F, aldehyde 8F, and the protected aldehydes 8A, 8B, and 8C.

We prepared a number of symmetrical esters 2 in which the catechol ring or replacement was the same on both the alcohol and acid moieties via DCC coupling of the acids 7 and alcohols 5 (Scheme 1). Each of these esters was then subjected to Pd-catalyzed decarboxylative coupling to afford protected rooperol 1′AA and the protected symmetrical rooperol derivatives 1′BB and 1′CC as well as the rooperol analogues 1DD, 1EE, and 1FF (Scheme 1).

In addition to these symmetrical rooperol analogues, two analogues with two different replacements for the catechol moieties were also prepared (Scheme 3). DCC coupling of the alcohol 4E with phenylpropiolic acid afforded the ester 2EG, while coupling with the acid 7A afforded the ester 2EA. Each of these was subjected to Pd-catalyzed decarboxylative coupling to afford the rooperol analog 1EG and the protected rooperol analog 1′EA (Scheme 3).

Scheme 3

Synthesis of unsymmetrical rooperol analogs and deprotection reactions.

The protected rooperol analogues 1′ were deprotected by two different methods. For the analogs containing a catechol group, we used our previously described silyl deprotection method [14] using HBr and KF, which afforded rooperol (1AA) and analogue 1EA (Scheme 3). The other analogues were prepared by AcOH-buffered TBAF deprotection to afford 1BB and 1CC (Scheme 3).

With these rooperol analogues in hand, we evaluated the utility of the various catechol replacements by determining the cancer cell cytotoxicity of these compounds versus rooperol using a MTT assay (Table 1). Rooperol displays cytotoxicity against all three cell lines examined: HeLa (cervical adenocarcinoma), H460 (lung carcinoma), and A549 (lung carcinoma). All of the rooperol analogues examined here were also tested against HeLa cells, which were slightly more sensitive to rooperol compared to the other cell lines. All of the analogues, with the exception of 1DD, show some activity against HeLa cells, with analogues 1CC and 1EA displaying activity close to that of rooperol. Interestingly, while the symmetrical analogue 1EE lacks good activity against HeLa cells, the two asymmetrical analogues containing the same methylenedioxyphenyl catechol replacement, 1EA and 1EB, show better cytotoxicity. It is also interesting that 1EE has nearly the same activity as rooperol against A549 cells, while 1DD is much less active against both A549 and H460 cells compared to rooperol.

Table 1

Cancer Cell Cytotoxicity of Rooperol and Analogues.

Compound	HeLa (GI50, µM)	H460 (GI50, µM)	A549 (GI50, µM)
Rooperol (1AA)	18 ± 2	18.8 ± 0.7	26.0 ± 0.1
1BB	33.2 ± 0.7	nd	nd
1CC	77 ± 10	nd	nd
1DD	>500	221.8 ± 0.1	291.7 ± 0.1
1EE	112 ± 3	61.5 ± 0.1	28.4 ± 0.1
1FF	217 ± 16	nd	nd
1EA	38 ± 8	nd	nd
1EG	77± 12	nd	nd

3. Discussion

A scalable and general synthesis of rooperol and analogues has been developed and used to evaluate the cancer cell cytotoxicity of a number of analogues that may have improved metabolic stability compared to rooperol. The synthesis of rooperol presented here is longer (seven total steps, longest linear sequence of five steps) but higher yielding (27% vs. 17%) compared to our previous total synthesis [14]. In addition to preparing rooperol, this route was employed to prepare a series of symmetrical and unsymmetrical analogues that were evaluated for cancer cell cytotoxicity compared to rooperol. The symmetrical compound 1BB, bearing 4-hydroxyl substituents on each aromatic ring, displays activity against HeLa cells that is approximately one-half that of rooperol (GI50 = 33.2 vs. 18 µM) and similar to an unsymmetrical analog bearing a methylenedioxyphenyl group (1EA) in place of one catechol group of rooperol. Interestingly, while the symmetrical analogue bearing two methylenedioxyphenyl groups, 1EE, is much less active against HeLa cells compared to rooperol, 1EE is very similar to rooperol in activity against A459 cells (GI50 = 28.4 vs. 26.0 µM). Notably, because 1EE lacks the catechol moieties of rooperol, it is not as prone to redox cycling and therefore chemically more stable than rooperol. Together, these results indicate that a more extensive search for symmetrical and asymmetrical rooperol analogues is warranted. In addition, the results obtained here indicate that the phenol and methylenedioxyphenyl catechol replacements are promising, the latter particularly against lung cancer cells. Work establishing the metabolic stability of these analogues versus rooperol is on-going.

4. Materials and Methods

All reactions were carried out under argon in oven-dried glassware with magnetic stirring. Unless otherwise noted, all materials were obtained from commercial suppliers and were used without further purification. THF was distilled from sodium/benzophenone prior to use. Flash chromatography was performed with EM Reagent silica gel (230–400 mesh) using the mobile phase indicated. Melting points (open capillary) are uncorrected. Unless otherwise noted, 1H and 13C NMR spectra were determined in CDCl3 on a spectrometer operating at 400 and 100 MHz, respectively, and are reported in ppm using solvent as internal standard (7.26 ppm for 1H and 77.0 ppm for 13C in CDCl3). Mass spectra were obtained by atmospheric pressure chemical ionization (APCI), chemical ionization using methane as the ionizing gas (CI), or by electrospray ionization (ESI). Copies of all NMR and MS spectra are available in the Supporting Information.

General Procedure-Silyl Protection of Benzaldehydes: 1,2-bis-(()oxy)benzaldehyde (8A) [17]: To solution of 3,4-dihydroxybenzaldehyde (4 g, 29 mmol) in dichloromethane (120 mL) under argon was added imidazole (7.9 g, 4 equivalent). The reaction mixture was cooled in an ice bath and tert-butyldimethyl-silyl chloride (12 g, 2.5 equivalent) was added. The reaction mixture was allowed to stir overnight, then diluted with dichloromethane and washed twice with 1 N HCl, once with saturated aqueous NaHCO3, once with brine, and then, the organic layer was dried over Na2SO4. After filtration, the solution was evaporated under vacuum and the residue subjected to flash chromatography purification (SiO2, 10% EtOAc/ hexanes) to yield the product (9.7 g, 91%) as a viscous oil, which crystallized to a pale yellow solid. Mp: 42–43 °C (lit: 39–41 °C [18]); IR (neat, ATR): cm−1 2953, 2926, 2856, 1694, 1569, 1504, 1298, 1284, 1269, 1248, 1213, 1157, 1104, 973, 897, 825, 781, 731; 1H NMR (400 MHz, CDCl3): δ 9.81 (s, 1H), 7.38–7.34 (m, 2H), 6.94 (d, 1H, J = 8.0 Hz), 0.99 (s, 18H), 0.25 (s, 6H), 0.23 (s, 6H); 13C{1H} NMR (100 MHz, CDCl3): δ 190.7, 153.3, 147.6, 130.7, 125.2, 120.7, 120.5, 25.8, 25.7, 18.4, 18.2, −4.0, −4.2; MS (APCI, pos) m/z (%): 367(67) [M + H]+.

4-(()oxy)benzaldehyde (8B) [19]: Prepared from 4-hydroxybenzaldehyde following the general procedure described for 8A, which afforded a quantitative yield (9.69 g) of a colorless oil. IR (neat, ATR): cm−1 2955, 2930, 2858, 1697, 1506, 1255, 1154, 902; 1H NMR (500 MHz, CDCl3): δ 9.89 (s, 1H), 7.79 (d, 2H, J = 8.6 Hz), 6.94 (d, 2H, J = 8.6 Hz), 0.99 (s, 9H), 0.25 (s, 6H); 13C{1H} NMR (125 MHz, CDCl3): δ 190.8, 161.5, 131.9, 130.4, 120.5, 25.5, 18.2, −4.3; MS (ESI, pos) m/z (%): 237(55) [M + H]+.

3,5-bis(()oxy)benzaldehyde (8C) [20]: Prepared from 3,5-dihydroxybenzaldehyde following the general procedures described above for 8A, which afforded a 93% yield (1.28 g) of a slightly yellow oil that slowly crystalized in the freezer. Mp = 28–29 °C; IR (neat, ATR, cm−1) 2955, 2930, 2857, 1702, 1587, 1330, 1253, 1165, 826; 1H NMR (500 MHz, CDCl3): δ 9.86 (s, 1H), 6.95 (d, 2H, J = 2.3 Hz), 6.59 (t, 1H, J = 2.3 Hz), 0.99 (s, 18H), 0.22 (s, 12H); 13C{1H} NMR (125 MHz, CDCl3): δ 191.7, 157.3, 138.4, 118.4, 114.3, 25.6, 18.2, −4.4; MS (APCI, pos) m/z (%): 367(17) [M + H]+.

2,2-dimethyl-4(F): Prepared as previously reported [21] from 4-bromosalicyl alcohol isopropylidene acetal to afford 11.17 g (93% yield) of colorless oil that slowly recrystallized in the freezer to afford off-white crystals. Mp = 55–58 °C (lit. 56–58 °C [21]); IR (neat, ATR): cm−1 2992, 2869, 1690, 1496, 1384, 1269; 1H NMR (500 MHz, CDCl3): δ 9.85 (s, 1H), 7.70 (dd, 1H, J = 8.5, 1.0 Hz), 7.55 (t, 1H, J = 1.0 Hz), 6.93 (d, 1H, J = 8.5 Hz), 4.90 (s, 2H), 1.57 (s, 6H); 13C{1H} NMR (125 MHz, CDCl3): δ 190.7, 156.8, 130.5, 129.4, 126.6, 119.7, 117.7, 100.8, 60.6, 24.8; MS (ESI, pos) m/z (%): 215(14) [M + Na]+, 191(15), 161(11).

General Procedure—Wittig reaction: Methyl ()-3-(3,4-bis(()oxy)phenyl)acrylate (4A) [22]: To a solution of methyl (triphenylphosphoranylidene)acetate (18.4 g, 55 mmol) in 200 mL of DCM at room temperature was added a ca. 1 M solution of 3,4-bis(tert-butyldimethylsilyloxy)benzaldehyde (8A, 6.33 g, 45.8 mmol) in DCM dropwise over 5 min. Upon completion of the addition, the mixture was stirred for an additional 18 h at room-temperature, and the solvent was removed by evaporation. The resulting pasty oil was diluted with 40 mL of hexanes, and the Ph3PO that precipitated was removed by filtration, and the filter cake was washed with two 40 mL portions of hexanes. The combined organic layers were concentrated under reduced pressure. Flash chromatography on silica gel (11:1 hexanes/EtOAc) of the residue afforded 4.17 g (91%) of 4′A0 as a white solid. Mp = 63.0–64.0 °C; IR (ATR, neat): cm−1 2947, 2930, 2857, 1721, 1631, 1506, 1472, 1422, 1289, 1249, 1161, 1126, 911, 838, 813, 777; 1H NMR (500 MHz, CDCl3): δ 6.72 (d, 2H, J = 2.1 Hz), 7.64 (d, 1H, J = 15.9 Hz), 6.46 (t, 1H, J = 2.1 Hz), 6.44 (d, 1H, J = 15.9 Hz), 3.88 (s, 3H), 0.99 (s, 18H), 0.19 (s, 12H); 13C{1H} NMR (125 MHz, CDCl3): δ 167.7, 149.4, 147.1, 144.8, 128.0, 122.2, 121.1, 120.4, 115.4, 51.5, 25.9, 25.8, 18.5, 18.4, −4.0, −4.1; MS (APCI, pos) m/z (%): 423(66) [M + H]+.

Methyl ()-3-(4-(()oxy)phenyl)acrylate (4B) [23]: Prepared from aldehyde 8B (2.22 g, 9.4 mmol) following the general procedure described for 4A, which afforded a 4B as a white solid (2.73 g, 91% yield). Mp = 34.0–36.0 °C; IR (ATR, neat): cm−1 2953, 2928, 2856, 1710, 1635, 1598, 1508, 1436, 1324, 1251, 1192, 1166, 988, 908, 834, 780; 1H NMR (400 MHz, CDCl3): δ 7.64 (d, 1H, J = 15.9), 7.41 (d, 2H, J = 8.6), 6.84 (d, 3H, J = 8.6), 6.30 (d, 1H, J = 15.9), 3.79 (s, 3H), 0.99 (s, 9H), 0.21 (s, 6H); 13C{1H} NMR (100 MHz, CDCl3): δ 167.6, 157.8, 144.5, 129.6, 127.6, 120.4, 120.4, 115.4, 51.5, 25.6, 18.2, −4.4; MS (ESI, pos) m/z (%): 293(24) [M + H]+.

Methyl ()-3-(3,5-bis(()oxy)phenyl)acrylate (4C) [24]: Prepared from aldehyde 8C following the general procedure described for 4A, which afforded 4C as a colorless oil, (3.93 g, 95% yield). IR (ATR, neat) cm−1 2951, 2928, 2857, 1716, 1638, 1578, 1437, 1281, 1154, 1002, 828, 778; 1H NMR (500 MHz, CDCl3): δ 7.55 (d, 1H, J = 15.9 Hz), 6.62 (dd, 2H, J = 2.2, 0.4 Hz), 6.36 (t, 1H, J = 2.2 Hz), 6.34 (d, 1H, J = 15.9 Hz), 3.80 (s, 3H), 0.98 (s, 18H), 0.20 (s, 12H); 13C{1H} NMR (125 MHz, CDCl3): δ 167.4, 156.9, 144.8, 136.1, 117.9, 114.2, 113.2, 51.7, 25.6, 18.2, −4.4; MS (APCI, pos) m/z (%): 423(58) [M + H]+.

Methyl ()-3-(3,4-difluorophenyl)acrylate (4D) [25]: Prepared from 3,4-difluorobenzaldehyde following the general procedure described for 4A, which afforded 4D as a white solid (3.17 g, 91%). Mp = 76.9–77.9 °C; IR (ATR, neat): cm−1 3057, 2957, 2920, 1704, 1640, 1514, 1494, 1435, 1330, 1270, 1250, 1220, 1189, 1175, 1144, 1111, 987, 870, 814, 790; 1H NMR (400 MHz, CDCl3): δ 7.57 (d, 1H, J = 16.0 Hz), 7.32 (ddd, 1H, J = 11.0, 7.6, 2.1 Hz), 7.25–7.21 (m, 1H), 7.20–7.12 (m, 1H), 6.33 (dd, 1H, J = 16.0, 0.3 Hz), 3.79 (s, 3H); 13C{1H} NMR (100 MHz, CDCl3): δ 166.8, 151.5 (dd, J = 253, 13 Hz), 153.4 (dd, J = 249, 13 Hz), 142.4, 131.6 (dd, J = 5, 5 Hz), 124.7 (dd, J = 6, 3 Hz), 118.9 (d, J = 2 Hz), 117.8 (d, J = 17 Hz), 116.3 (d, J = 17 Hz), 51.7; 19F{1H} NMR (376 MHz, CDCl3): δ −134.2 (d, J = 21 Hz), −136.6 (d, J = 21 Hz); MS (APCI, pos) m/z (%): 199(5) [M + H]+.

Methyl ()-3-(benzo[[1,3]dioxol-5-yl)acrylate (4E) [26]: Prepared from piperonal following the general procedure described for 4A, which afforded 4E as a white solid (5.4 g, 96%). Mp: 133–135 °C; IR (ATR, neat): cm−1 2950, 2901, 1699, 1622, 1597, 1594, 1495, 1454, 1438, 1367, 1304, 1255, 1201, 1169, 1124, 1105, 1035, 1004, 931, 916, 821; 1H NMR (400 MHz, CDCl3): δ 7.53 (d, 1H, J = 16.0 Hz), 6.98–7.04 (m, 2H), 6.82 (d, 1H, J = 8.4 Hz), 6.26 (d, 1H, J = 16.0 Hz), 6.01 (s, 2H), 3.79 (s, 3H); 13C{1H} NMR (100 MHz, CDCl3): δ 167.6, 149.6, 148.3, 144.5, 128.8, 124.4, 115.7, 108.5, 106.4, 101.5, 51.6; MS (ESI, pos) m/z (%): 207(78) [M + H]+.

Methyl ()-3-(2,2-dimethyl-4[1,3]dioxin-6-yl)acrylate (4F): Prepared from 2,2-dimethyl-4H-benzo(d)(1,3)dioxine-6-carbaldehyde (8F) following the general procedure described for 4A, which afforded 4F as a white, waxy solid (249 mg, 99%). IR (ATR, neat): cm−1 2992, 2947, 2860, 1716, 1627, 1609, 1582, 1497, 1448, 1463, 1384, 1374, 1267, 1191, 1166, 1114, 1062, 953, 863, 830, 796; 1H NMR (400 MHz, CDCl3): δ 7.60 (1H, dd, J = 16.0, 3.0 Hz), 7.33–7.35 (m, 1H), 7.13 (s, 1H), 6.81 (dd, 1H, J = 8.4, 3.3 Hz), 6.26–6.29 (m, 1H), 4.83 (s, 2H), 3.77 (s, 3H), 1.53 (s, 6H); 13C{1H} NMR (100 MHz, CDCl3): δ 167.6, 153.3, 144.4, 128.0, 126.7, 124.9, 119.7, 117.6, 115.4, 100.1, 60.6, 51.5, 24.7; MS (ESI, pos) m/z (%): 271(10) [M + Na]+, 304(71) [M + Na + CH3OH]+; HRMS (ESI, pos): m/z calculated for C14H16O4 [M + H]+ 249.1121, found 249.1124.

General Procedure–DIBAL-H Reduction: ()-3-(3,4-bis(()oxy)phenyl)prop-2-en-1-ol (5A): To a solution of 4A (3.0 g, 7.1 mmol) in 70 mL of dry DCM in a 250 mL rb flask at −78 °C under Ar was added dropwise a solution of DIBAL-H (1.2 M solution in hexane, 18 mL, 21.6 mmol). Upon completion of the addition, the reaction mixture was allowed to warm to room temperature over 2 h and then carefully added dropwise to a stirred mixture of 2 M HCl (100 mL) and ice. After stirring for 30 min, the aqueous layer was extracted with CH2Cl2 (3 × 100 mL). The combined organic layers were washed with brine, dried over Na2SO4, and concentrated under reduced pressure to afford a colorless oil that was subjected to flash chromatography purification (SiO2, 0–10% EtOAc/hexanes) to yield the product (2.25 g, 80%) as a colorless oil. IR (ATR, neat): cm−1 3360(br), 2929, 2886, 2857, 1598, 1511, 1471, 1419, 1301, 1254, 1124, 989, 905, 840, 781; 1H NMR (400 MHz, CDCl3): δ 6.88 (d, 1H, J = 2.2 Hz), 6.85 (dd, 1H, J = 8.3, 2.2 Hz), 6.77 (d, 1H, J = 8.2 Hz), 6.48 (dt, 1H, J = 15.8, 1.3 Hz), 6.18 (dt, 1H, J = 15.8, 5.9 Hz), 4.30–4.27 (m, 1H), 1.38 (t, 1H, J = 5.7 Hz), 0.99 (s. 9H), 0.98 (s, 9H), 0.20 (s, 6H), 0.19 (s, 6H); 13C{1H} NMR (100 MHz, CDCl3): δ 146.9, 131.2, 130.2, 126.3, 121.0, 119.9, 119.0, 63.9, 25.9, 25.8, 18.5, 18.4, −4.1; MS (ESI, pos) m/z (%): 417(25) [M + Na]+, 377(15); HRMS (ESI, pos): m/z calculated for C21H38NaO3Si2 [M + Na]+ 417.2257, found 417.2253.

()-3-(4-(()oxy)phenyl)prop-2-en-1-ol (5B) [23]: Prepared from 4B following the general procedure described for 5A, which afforded 5B as a colorless oil, 1.3 g (85%). IR (ATR, neat): cm−1 3325(br), 2954, 2928, 2857, 2602, 1507, 1252, 1168, 908, 835, 799, 778; 1H NMR (400 MHz, CDCl3): δ 7.26 (d, 2H, J = 8.5 Hz), 6.79 (d, 2H, J = 8.5 Hz), 6.55 (d, 1H, J = 15.9 Hz), 6.24 (dt, 1H, J = 15.5, 5.9 Hz), 4.39 (dd, 2H, J = 5.9, 1.2 Hz), 2.36 (d, 1H, J = 0.3 Hz), 0.98 (s, 9H), 0.20 (s, 6H); 13C{1H} NMR (100 MHz, CDCl3): δ 155.5, 131.0, 129.9, 127.6, 126.4, 120.2, 63.9, 25.7, 18.2, -4.4; MS (APCI, pos) m/z (%): 264(5) [M]+, 247(67).

()-3-(3,5-bis(()oxy)phenyl)prop-2-en-1-ol (5C) [24]. Prepared from 4C following the general procedure described for 5A, which afforded 5C as a colorless oil, 914 mg (95%). IR (neat, ATR): cm−1 3292(br), 2928, 2857, 1581, 1435, 1251, 1164, 1021, 827, 777; 1H NMR (400 MHz, CDCl3): δ 6.50–6.46 (m, 3H), 6.29 (dt, 1H, J = 10.0, 5.0 Hz), 6.25 (t, 1H, J= 2.1 Hz), 4.29 (dd, 2H, J = 5.6, 1.3Hz), 0.98 (s, 18H), 0.19 (s, 12H); 13C{1H} NMR (100 MHz, CDCl3): δ 156.6, 138.4, 131.0, 128.6, 111.7, 111.6, 63.7, 25.7, 18.2, −4.4.

()-3-(3,4-difluorophenyl)prop-2-en-1-ol (5D). Prepared from 4D following the general procedure described for 5A, which afforded 5D as a colorless oil, 1.18 g (92%). IR (neat, ATR): cm−1 3271(br), 2987, 2845, 1603, 1514, 1289, 1270, 1090, 1018, 967; 1H NMR (400 MHz, CDCl3): δ 7.21–7.16 (m, 1H), 7.13–7.06 (m, 2H), 6.53 (d, 1H, J = 15.9 Hz), 6.27 (dt, 1H, J = 15.9, 5.5 Hz), 4.32 (dd, 2H, J = 5.5, 1.4 Hz); 13C{1H} NMR (100 MHz, CDCl3): δ 150.5 (dd, J = 247, 13 Hz), 149,8 (dd, J = 249, 13 Hz), 133.9 (dd, J = 6, 4 Hz), 129.6 (d, J = 2 Hz), 128.8, 122.6 (dd, J = 6, 3 Hz) 117.3 (d, J = 18 Hz), 114.7 (d, J = 8 Hz), 63.2; 19F NMR (376 MHz, CDCl3) δ: −137.9 (d, J = 21 Hz), −139.0 (d, J = 21 Hz); MS (ESI, pos) m/z (%): 171 [M + H]+(9), 157(12); HRMS (EI, pos): m/z calculated for C9H8OF2 [M]+ 170.0543, found 170.0539.

()-3-(benzo[[1,3]dioxol-5-yl)prop-2-en-1-ol (5E). Prepared from 4E following the general procedure described for 5A, which afforded 5E as a white solid, 1.94 g (90%). Mp: 79.5–80.0 °C; IR (neat, ATR): cm−1 3350, 2920, 2895, 2851, 1499, 1441, 1243, 1083, 1034, 1003, 965, 920, 909; 1H NMR (500 MHz, CDCl3): δ 6.93 (d, 1H, J = 1.5 Hz), 6.81 (1H, dd, J= 8.0, 1.5 Hz), 6.75 (d, 1H, J = 8.0 Hz), 6.52 (dd, 1H, J = 15.8, 1.3 Hz) 6.20 (dd, 1H, J = 15.8, 5.9 Hz), 5.96 (s, 2H), 4.29 (td, 2H, J = 5.8, 1.3 Hz), 1.44 (t, 1H, J = 5.9 Hz); 13C{1H} NMR (100 MHz, CDCl3): δ 148.0, 147.3, 131.1, 131.0, 126.6, 121.1, 108.2, 105.7, 101.1, 63.7; MS (ESI, pos) m/z (%): 177(20) [M-H]+.

()-3-(2,2-dimethyl-4[1,3]dioxin-6-yl)prop-2-en-1-ol (5). Prepared from 4′C3 following the general procedure described for 4A0, which afforded a white solid, 248 mg (quant.). Mp: 130.0–130.9 °C; IR (neat, ATR): cm−1 3355(br), 2992, 2854, 1498, 1383, 1260, 1200, 1142, 1115, 1056, 1004, 975, 958, 868, 831; 1H NMR (500 MHz, CDCl3): δ 7.20 (dd, 1H, J = 8.5, 1.8 Hz), 6.98 (d, 1H, J = 2.0 Hz), 6.77 (d, 1H, J = 8.5 Hz), 6.49–6.52 (m, 1H), 6.21 (dt, 1H, J = 15.8, 5.9 Hz), 4.82 (s, 2H), 4.28 (dd, 2H, J = 5.9, 1.5 Hz), 1.54 (s, 6H); 13C{1H} NMR (125 MHz, CDCl3): δ 150.9, 130.8, 129.1, 125.5, 126.2, 122.7, 119.4, 117.3, 99.7, 63.8, 60.8, 24.7; MS (APCI, pos) m/z (%): 203(87) [M − H2O + H]+; HRMS (CI, pos) calculated for C13H16O3 [M]+ 220.1099, found 220.1101.

General Procedure–Vinyl Dibromide Formation: ((4-(2,2-dibromovinyl)-1,2-phenylene)bis(oxy))bis() (6A) [27]. To a 250 mL round-bottom flask with stirbar under Ar was placed 8.92 g (26.9 mmole) of CBr4. DCM (30 mL) was added to the flask via syringe, and the reaction flask was cooled in an ice bath. Triphenylphosphine (14.09 g, 53.7 mmole) was added the flask in portions, and the resulting mixture was stirred under Ar for 5 min. A solution of 4.94 g (13.5 mmole) of the aldehyde 8A in 10 mL DCM was added to the reaction mixture, and the ice bath was removed. The mixture was allowed to stir at room temperature for 30 min, when TLC monitoring indicated the reaction was complete. Saturated sodium bicarbonate solution was carefully added until the aqueous layer was neutral by pH paper. The aqueous layer was then extracted with three 80 mL portions of DCM. The combined organic extracts were washed with saturated brine and then dried over anhydrous sodium sulfate. The dried solution was filtered and then evaporated under reduced pressure, and the residue subjected to chromatography (0–5% EtOAc/hex) to afford 6.36 g (91 % yield) of 6A as a slightly yellow oil. IR (ATR, neat): cm−1 2952, 2928, 2857, 1503, 1471, 1294, 1251, 1128, 881, 903, 835, 777; 1H NMR (400 MHz, CDCl3): δ 7.38 (s, 1H), 7.28 (d, 1H, J = 2.3 Hz), 6.99 (ddd, 1H, J = 8.3, 2.3, 0.6 Hz), 6.85 (d, 1H, J = 8.3 Hz), 1.05 (s, 9H), 1.04 (s, 9H), 0.28 (s, 6H), 0.27 (s, 6H); 13C{1H} NMR (100 MHz, CHCl3): δ 147.5, 145.6, 136.4, 128.4, 122.5, 120.7, 120.6, 86.8, 25.9, 25.8, 18.5, 18.4, −4.1; MS (ESI, neg) m/z (%): 521(2), 519(1), 523(1) [M − H]–.

(4-(2,2-dibromovinyl)phenoxy)dimethylsilane (6B): Prepared from 8B following the general procedure described for 6A to afford 2.26 g (58% yield) of 8B as a colorless oil: IR (ATR, neat): cm−1 2953, 2928, 2856, 1601, 1505, 1462, 1252, 1171, 908, 870, 836, 775, 691; 1H NMR (500 MHz, CDCl3): δ 7.44–7.46 (m, 2H), 7.40 (s, 1H), 6.82–6.83 (m, 2H), 0.99 (s, 9H), 0.22 (s, 6H); 13C{1H} NMR (125 MHz, CHCl3): δ 156.0, 136.4, 129.8, 128.3, 119.9, 87.2, 25.6, 18.2, −4.4; MS (ESI, neg) m/z (%):393(8), 391(15), 389(7) [M − H] –; HRMS (EI, pos): m/z calculated for C14H20OSiBr2 [M]+ 391.9630, found 391.9636.

((5-(2,2-dibromovinyl)-1,3-phenylene)bis(oxy))bis() (6C): Prepared from 8C following the general procedure described for 6A to afford 3.83 g (89% yield) of 6C as a pale yellow oil. IR (ATR, neat): cm−1 2854, 2929, 2857, 1580, 1431, 1334, 1252, 1163, 1030, 827, 810, 338; 1H NMR (400 MHz, CDCl3) δ: 7.53 (2, 1H), 6.65 (dd, 2H, J = 2.1, 0.4 Hz), 6.33 (t, 1H, J= 2.1 Hz), 0.98 (s, 18H), 0.20 (s, 12H); 13C{1H} NMR (100 MHz, CDCl3) δ 156.5, 136.7, 136.6, 113.4, 112.7, 89.4, 25.7, 18.2, −4.38; MS (ESI, neg) m/z (%): 521(6) [M − H]-; HRMS (EI, pos): m/z calculated for C20H34O2Si2Br2 (M)+ 522.0444, found 522.0441.

4-(2,2-dibromovinyl)-1,2-difluorobenzene (6D) [28] Prepared from 3,4-difluorobenzaldehyde following the general procedure described for 6A to afford 10.13 g (97% yield) of a 6D as a colorless oil. IR (neat, ATR): cm−1 3019, 1604, 1513, 1432, 1418, 1287, 1214, 1115, 882; 1H NMR (400 MHz, CDCl3): δ 7.22–7.15 (m, 1H), 7.39 (d, 1H, J = 0.27), 7.39–7.48 (m, 2H); 13C{1H} NMR (100 MHz, CDCl3): δ 91.4 (d, J = 2.52), 117.1 (d, J = 16.2 Hz), 117.3 (d, J = 14.8 Hz), 125.5 (dd, J = 6.5, 3.6), 132.1 (dd, J = 6.5, 4.3), 135.1 (d, J = 1.6), 150.0 (dd, J = 247.9, 11.7 Hz), 150.1 (dd, J = 249.9, 10.7 Hz); 19F{1H} NMR (376 MHz, CDCl3) δ: 136.2 (d, J = 20.7 Hz), 136.8 (d, J = 20.7 Hz).

5-(2,2-dibromovinyl)benzo( (6E) [29]: Prepared from piperonal following the general procedure described for 6A to afford 3.87 g (95% yield) of 6E as a crystalline yellow solid. Mp = 50.9–51.7 °C; IR (ATR, neat): 2962, 2907, 1500, 1486, 1443, 1305, 1255, 1192, 1098, 1032, 924, 864, 840, 798, 752; 1H NMR (400 MHz, CDCl3) δ: 5.98 (2H, s), 6.80 (1H, d, J = 8.1 Hz), 6.95 (1H, ddd, J = 8.1, 1.7, 0.7 Hz), 7.18 (1H, dd, J = 1.7, 0.4), 7.37 (1H, s); 13C{1H} NMR 100 MHz, CDCl3) δ: 87.9, 101.4, 108.1, 108.2, 123.4, 129.2, 136.3, 147.6, 147.8; MS (APCI, pos) m/z (%): 309(1), 307(2), 305(1) [M + H]+.

General Procedure–Corey-Fuchs Elimination with Trapping by CO(3,4-bis(()oxy)phenyl)propiolic acid (7A): To a solution of 5.0 g (9.57 mmol) of the vinyl dibromide 6A in 40 mL of THF under argon at −78 °C was added dropwise 8.8 mL of a 2.6 M solution (2.4 equivalent) of n-butyllithium in hexane. The reaction mixture was stirred at −78 °C for 1h, the ice bath was removed, and the reaction mixture was allowed to warm to room temperature. Chips of clean dry ice (ca. 5 g, large excess) were carefully added to the reaction mixture and allowed to fully dissolve/sublime while the mixture returned to room temperature. Water (20 mL) was added to the reaction mixture, and the THF was removed under reduced pressure. The residue was transferred to a separatory funnel, and ice-cold 1 M HCl (75 mL) and DCM (100 mL) were added. The aqueous phase was acidified with 6N HCl until the organic phase was no longer cloudy. The organic phase was collected. The aqueous phase was extracted twice more with 50 mL DCM, and extracts were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and evaporated. Recrystallization (EtOAc/hex) of the residue afford 2.77g (78%) of 7A as a white solid. Mp = 143–145 °C. IR (KBr): cm−1 3100, 2955, 2932, 2859, 2208, 1676, 1507; 1H NMR (400 MHz, CDCl3): δ 7.13 (1H, dd, J = 8.3, 2.0 Hz), 7.07 (1H, d, J = 2.0 Hz), 6.81 (1H, d, J = 8.3 Hz), 0.99 (9H, s), 0.98 (9H, s), 0.22 (6H, s), 0.21 (6H, s). 13C{1H} NMR (100 MHz, CDCl3): δ 158.5, 150.8, 147.0, 127.9, 125.7, 121.2, 111.4, 90.1, 79.3, 25.8, 18.5, 18.4, −4.1, −4.2; MS (ESI, neg) m/z (%): 405(18) [M − H]–; Matches lit [14].

3-(4-(()oxy)phenyl)propiolic acid (7B). Prepared from 6B following the general procedure described for 7A to afford 560 mg (62% yield) of 7B as a tan solid. Mp = 104.2–106.3 °C; IR (ATR, neat): 2951, 2928, 2857, 2197, 1667, 1595, 1507, 1255, 1208m 1163, 903, 835, 781 cm−1; 1H NMR (400 MHz, CDCl3) δ: 7.51 (d, 2H, J = 8.7 Hz), 6.83 (d, 2H, J = 8.7 Hz), 0.98 (s, 9H), 0.22 (s, 6H). 13C{1H} NMR (100 MHz, CDCl3): δ 158.8, 158.6, 135.3, 120.5, 111.5, 90.0, 79.8, 25.5, 18.2, −4.4; MS (ESI, neg) m/z (%): 275(15) [M − H]–, 231(46), 160(6), 135(16), 117(7); HRMS (ESI, neg): m/z calculated for C15H20O3Si [M − H]– 275.1109, found 275.1112.

3-(3,5-bis(()oxy)phenyl)propiolic acid (7C): Prepared from 6C following the general procedure described for 7A to afford 1.6 g (quant. yield) of 7C as a white semi-solid. IR (ATR, neat): cm−1 2954, 2928, 2857, 2223, 1677, 1578, 1427, 1251, 1167, 1028, 825, 776; 1H-NMR (CDCl3, 500 MHz): δ 6.71 (2H, d, J = 2.2 Hz), 6.45 (1H, t, J = 2.2 Hz), 0.97 (18H, s), 0.21 (12H, s); 13C{1H} NMR (100 MHz, CHCl3): δ 158.2, 156.7, 120.0, 118.1, 116.1, 88.9, 79.4, 25.6, 18.2, −4.4; MS (ESI, neg) m/z (%): 405(11) (M-H), 361(49), 247(13),101(8); HRMS (ESI, neg): m/z calculated for C9H4F2O2 [M − H]– 181.0107, found 181,0110.

3-(3,4-difluorophenyl)propiolic acid (7D). Prepared from 6D following the general procedure described for 7A to afford 193 mg (23% yield) of 7D as a pale yellow solid. Mp 169.0–171.6 °C; IR (ATR, neat): cm−1 3078, 2901, 2851, 2221, 1795, 1685, 1629, 1601, 1511, 1438, 1277, 1251, 1207, 1042, 916, 865; 1H-NMR (CDCl3, 400 MHz): δ 10.18 (s(br), 1H), 7.46–7.42 (m, 1H), 7.40–7.39 (m, 1H), 7.38 (q, 1H, J = 9.0 Hz); 13C{1H} NMR (100 MHz, CHCl3): δ 167.7, 151.6 (dd, J = 253, 12 Hz), 149.8 (dd, J = 248, 13 Hz), 130.6, 121.8 (d, J = 19 Hz), 118.6 (d, J = 18 Hz), 116.7 (d, J = 18 Hz), 116.3 (dd, J = 4, 3 Hz), 82.1 (d, J = 3 Hz), 82.0; MS (ESI, neg) m/z (%): 181(21) (M-H); HRMS (ESI, neg): m/z calculated for C9H4F2O2 [M − H]– 181.0107, found 181.0110.

3-(benzo()propiolic acid (7E) [30]. Prepared from 6E following the general procedure described for 7A to afford 1.9 g (81% yield) of 7E as an orange solid. Mp = 162.8–165.6 °C (dec); IR (ATR, neat): cm−1 2982, 2915, 2871, 2206, 1667, 1489, 1446, 1410, 1305, 1243, 1193, 1098, 1035, 923, 860, 811 cm−1;1H NMR (400 MHz, CD3OD) δ: 6.02 (2H, s); 6.86 (1H, dd, J = 8.06, 0.36), 7.01 (1H, dd, J = 1.61, 0.35), 7.15 (1H, dd, J = 8.06, 1.64); 13C NMR (400 MHz, CD3OD) δ: 80.7, 87.2, 103.3, 109.8, 113.1, 113.8, 129.7, 149.3, 151.6, 156.8; MS (ESI, neg) m/z (%): 189(9) [M − H]–, 145(14), 75(54).

3-(2,2-dimethyl-4)propiolic acid (7F): A solution of 300 mg (1.6 mmole) of 6-ethynyl-2,2-dimethyl-4H-benzo(d)(1,3)dioxine [16] in 1 mL of THF was cooled to −78 °C. To this solution was added dropwise 637 µL (1.6 mmole) of 2.5 M BuLi in hexanes. The solution was warmed to −20 °C, stirred at −20 °C for 30 min, and then, CO2 was bubbled through the reaction mixture over 4 h during which the reaction was allowed to warm to rt. The reaction was quenched by addition of a small amount of water, the solvent was evaporated, and the residue was subjected to flash chromatography (10% EtOAc/Hex + 1% AcOH) to afford 229 mg (62%) of 7F as a white solid. Mp = 119.5–121.0 °C; IR (ATR, neat): cm−1 2989, 2938, 2866, 2205, 1659, 1608, 1495, 1327, 1273, 1252, 1218, 1120, 884; 1H-NMR (CDCl3, 500 MHz): δ 7.32 (1H, dd, J = 8.8, 2.0 Hz), 7.28 (1H, d, J = 2.0 Hz), 6.81 (1H, d, J = 8.5 Hz), 4.83 (2H, s), 1.55 (6H, s); 13C{1H} NMR (125 MHz, CHCl3): δ 157.6, 154.0, 144.6, 130.4, 119.9, 117.8, 110.6, 100.6, 89.6, 79.5, 60.4, 24.8; MS (ESI, neg) m/z (%): 231(4) [M − H]–, 187(38), 129(11), 75(4), 69(14), 59(20); HRMS (ESI, neg): m/z calculated for C13H12O4 [M − H]- 231.0663, found 231.0665.

General Procedure – DCC Coupling: ()-3-(3,4-bis(()oxy)phenyl)allyl 3-(3,4-bis(()oxy)phenyl)propiolate (2AA): The carboxylic acid (7A) (2.413 g, 5.93 mmol) and DMAP (112 mg, 0.92 mmol) were placed in a round-bottomed flask that was then purged with argon. To the flask was added 40 mL of dry DCM and ca. 1 g of 4Å molecular sieves. A solution of the alcohol (4A) (1.815 g, 4.60 mmol) in 15 mL DCM was added via syringe. The flask was placed in an ice bath, and a solution of 1.43g (6.90 mmol) of DCC in 3 mL DCM was added dropwise via syringe. The ice bath was removed, and the reaction was allowed to stir at room temperature for 2 h. The crude reaction mixture was filtered through a plug of silica gel, which was washed with 50 mL of 30% ethyl acetate/hexanes. The filtrate was evaporated and purified by flash chromatography (0–2% ethyl acetate/hexanes) to afford 2.91 g (81% yield) of ester 2AA as a pale yellow oil. IR (ATR, neat): cm−1 2955, 2930, 2216, 1711, 1511; 1H NMR (400 MHz, CDCl3): δ 7.10 (1H, dd, J = 8.3, 2.0 Hz), 7.04 (1H, d, J = 2.0 Hz), 6.88 (2H, m), 6.78 (2H, m), 6.59 (1H, d, J = 15.8 Hz), 6.14 (1H, dt, J = 15.8, 6.8 Hz), 4.84 (2H, dd, 6.8, 1.1 Hz), 0.99 (9H, s), 0.97 (18H, s), 0.96 (9H, s), 0.22 (6H, s), 0.21 (6H, s), 0.20 (6H, s), 0.19 (6H, s); 13C{1H} NMR (100 MHz, CD3COCD3): δ 154.1, 151.1, 148.0, 147.9, 147.7, 135.6, 131.1, 128.3, 126.0, 122.4, 122.0, 121.5, 121.3, 120.1, 112.9, 86.9, 80.5, 67.1, 26.4, 26.3, 26.2, 19.2, 19.09, 19.07, 19.04, −3.81, −3.84, −3.9; HRMS (ESI, pos) calc. for C42H70NaO6Si4 [M + Na]+ 805.4142, found 805.4148.

()-3-(4-(()oxy)phenyl)allyl 3-(4-(()oxy)phenyl)propiolate (2BB). Prepared from 4B and 7B following the general procedure described for 2AA to afford 64 mg (42% yield) of 2BB as a colorless oil. IR(ATR, neat): cm−1 2914, 1848, 2236, 1734, 1623, 1568, 1559, 1309, 1268, 1241, 1178, 1086, 640; 1H-NMR (CDCl3, 500 MHz): δ 7.48 (d, 2H, J = 8.8 Hz), 7.28 (d, 2H, J = 8.5 Hz), 6.83–6.79 (m, 4H), 6.65 (d, 1H, J = 15.8 Hz), 6.19 (dt, 1H, J = 15.8, 6.7 Hz), 4.85 (dd, 2H, J= 6.7, 1.1 Hz), 0.99 (s, 9H), 0.98 (s, 9H), 0.21 (s, 6H), 0.20 (s, 6H); 13C{1H} NMR (125 MHz, CDCl3): δ 158.1, 155.9, 154.1, 135.1, 134.9, 129.3, 127.9, 120.4, 120.2, 120.0, 112.0, 87.3, 80.1, 66.6, 25.6, 25.5, 18.2, −4.4; MS (ESI, pos) m/z (%): 523(3) [M + H]+.

()-3-(3,5-bis(()oxy)phenyl)allyl 3-(3,5-bis(()oxy)phenyl)propiolate (2CC): Prepared from 4C and 7C following the general procedure described for 2AA to afford 349 mg (70% yield) of 2CC as off-white needles. Mp = 77.4–80.7 °C; IR (ATR, neat): cm−1 2951, 1919, 1857, 2219, 1706, 1579, 1437, 1427, 1250, 1233, 1163, 1153, 1026, 825, 811, 777; 1H NMR (400 MHz, CD3Cl): δ 6.69 (d, 2H, J = 2.2 Hz), 6.59 (d, 1H, J = 15.8 Hz), 6.51 (d, 2H, J = 2.2 Hz), 6.43 (t, 1H, J = 2.2 Hz), 6.27–6.20 (m, 2H), 4.86 (dd, 1H, J = 6.6, 1.0 Hz), 0.98 (s, 18H), 0.97 (s, 18H), 0.19 (s, 12H), 0.18 (s, 12H); 13C{1H} NMR (100 MHz, CDCl3): δ 156.7, 156.6, 153.7, 137.7, 135.3, 122.1, 120.4, 177.9, 115.7, 112.2, 112.9, 86.6, 79.8, 66.5, 25.7, 25.6, 18.1, −4.4, −4.5; MS (ESI, pos) m/z (%): 783(9) (M + H), 395(11), 377(14); HRMS (APCI, pos) calculated for C42H70O6Si4 [M + H]+ 783.4322, found 783.4303.

()-3-(3,4-difluorophenyl)allyl 3-(3,4-difluorophenyl)propiolate (2DD): Prepared from 4D and 7D following the general procedure described for 2AA to afford 154 mg (76% yield) of 2DD as an amber oil. IR (ATR, neat): cm−1 2949, 2938, 2856, 2215, 1724, 1644, 1603, 1513, 1435, 1293, 1269, 1242, 1165, 1142, 965, 811, 785; 1H NMR (400 MHz, CDCl3): δ 7.42–7.33 (2H, m), 7.23–7.09 (4H, m), 6.63 (1H, d, J = 15.87), 6.23 (1H, dt, J = 15.84, 6.46), 4.87 (2H, dd, J = 6.47, 1.23); 13C{1H} NMR (100 MHz, CDCl3): δ 153.2, 152.1 (dd, J = 256, 13 Hz), 150.4 (dd, J = 251, 16 Hz), 150.2 (dd, J = 158, 23 Hz), 150.0 (dd, J = 251 13 Hz), 133.2 (dd, J = 6, 4 Hz), 133.1, 130.0 (dd, J = 7, 4 Hz), 123.1 (d, J = 2Hz), 123.0 (dd, J = 7, 4 Hz), 121.9 (d, J = 19 Hz), 118.0 (d, J = 17 Hz), 117.4 (d, J = 17 Hz), 116.2 (dd, J = 7, 4 Hz), 115.0 (d, J = 17 Hz), 84.1 (d, J = 2 Hz), 80.5, 66.1; 19F{1H} NMR (376 MHz, CDCl3) δ: −137.0 (d, J = 20 Hz), −139.3 (d, J = 21 Hz), −140.1 (d, J = 20 Hz), −140.9 (d, J = 21 Hz); MS (APCI, pos) m/z (%): 389(13) (M + MeOH + Na+); HRMS (CI, pos) calculated for C18H12F4O2 [M]+ 336.0773, found 336.0771.

()-3-(benzo()allyl 3-(benzo()propiolate (2EE) [31]: Prepared from 4E and 7E following the general procedure described for 2AA to afford 1.58 g (86% yield) of 2EE as an off-white, chalky solid. 86%. Mp = 104.9–106.5 °C; IR(ATR, neat): cm−1 2916, 2901, 2851, 2207, 1692, 1618, 1490, 1444, 1303, 1236, 1185, 1096, 1034, 931, 804; 1H NMR (400MHz, CDCl3): δ 7.15 (1H, dd, J = 8.1, 1.6), 7.00–6.93 (2H, m), 6.84 (1H, dd, J = 8.2, 1.6), 6.80–6.75 (2H, m), 6.62 (1H, d, J = 15.8), 6.15 (1H, d, J = 15.8), 5.95 (2, 2H), 5.94 (s, 2H), 4.83 (2H, dd, J = 6.7, 1.2); 13C{1H} NMR (100 MHz, CDCl3): δ 153.8, 150.0, 148.0, 147.7, 147.5, 135.0, 130.3, 128.8, 121.6, 120.2, 112.4, 108.7, 108.2, 105.8, 101.7, 101.1, 87.0, 79.4, 66.4. MS (ESI, pos) m/z (%): 373(3) [M + Na]+.

()-3-(2,2-dimethyl-4)allyl 3-(2,2-dimethyl-4)propiolate (2FF): Prepared from 4F and 7F following the general procedure described for 2AA to afford 15 mg (50% yield) of 2FF as a colorless oil. IR(ATR, CHCl3): cm−1 2993, 2940, 2856, 2211, 1704, 1612, 1580, 1497, 1384, 1375, 1312, 1290, 1270, 1254, 1234, 1198, 1144, 1115, 956, 873; 1H NMR (400 MHz, CDCl3): δ 7.39 (dd, 1H, J = 8.5, 1.9 Hz), 7.24–7.21 (m, 2H), 7.02 (s, 1H), 6.79 (t, 2H, J = 8.1 Hz), 6.62 (d, 1H, J = 15.8 Hz), 6.17 (dt, 1H, J = 15.8, 6.7), 4.85–4.81 (m, 4H), 4.80 (s, 2H), 1.54 (s, 12H); 13C{1H} NMR (100 MHz, CDCl3): δ 154.0, 153.6, 151.4, 134.9, 133.3, 130.1, 128.5, 126.6, 123.1, 120.2, 119.8, 119.4, 117.6, 117.3, 111.0, 100.4, 99.8, 87.1, 79.8, 66.6, 60.8, 60.4, 24.8, 24.7; MS (ESI, pos) m/z (%): 435(3) [M + H]+; HRMS (APCI, pos) calculated for C26H26O6 [M + H]+ 435.1802, found 435.1788.

()-3-(benzo()allyl 3-(3,4-bis(()oxy)phenyl)propiolate (2EA): Prepared from 4E and 7A following the general procedure described for 2AA to afford 139 mg (59% yield) of 2EA as slightly yellow needles. Mp 95.3–96.7 °C; IR(ATR, neat): cm−1 2927, 2855, 2208, 1556, 1493, 1437, 1411, 1318, 1275, 1238, 1155, 1119, 1035, 935, 896, 839, 783; 1H NMR (500MHz, CDCl3): δ 7.10 (dd, 1H, J = 8.3, 2.1 Hz), 7.05 (d, 1H, J = 2.1 Hz), 6.94 (d, 1H, J = 1.7 Hz), 6.84 (dd, 1H, J = 8.0, 1.7 Hz), 6.79 (d, 1H, J = 8.3 Hz), 6.76 (8.0 Hz), 6.63 (d, 1H, J = 15.8 Hz), 6.16 (dt, 1H, J = 15.8, 6.7 Hz), 5.96 (s, 2H), 4.84 (dd, 2H, J = 6.7, 1.2 Hz), 0.99 (s, 9H), 0.98 (s, 9H), 0.22 (s, 6H), 0.21 (s, 6H);13C{1H} NMR (125 MHz, CDCl3): δ 154.1, 150.2, 148.1, 147.7, 146.9, 135.1, 130.4, 127.4, 125.4, 121.6, 121.1, 120.3, 111.9, 108.2, 105.9, 101.1, 87.5, 79.6, 66.4, 25.8, 18.5, 18.4, −4.1, −4.2; MS (ESI, pos) m/z (%): 567(21) [M + H]+; HRMS (APCI, pos) calculated for C31H42O6Si2 [M + H]+ 567.2593, found 567.2586.

()-3-(benzo()allyl 3-phenylpropiolate (2EG) [30]: Prepared from 4E and phenylpropiolic acid following the general procedure described for 2AA to afford 179 mg (51% yield) of 2EG as a white solid. Mp = 55.7–57.2 °C. IR(ATR, neat) cm−1: 2996, 2896, 2210, 1697, 1501, 1490, 1444, 1299, 1280, 1251, 1182, 1170, 1125, 1036, 953, 930, 911, 752; 1H NMR (400MHz, CDCl3): δ 7.60–7.58 (m, 2H), 7.45 (t, 1H, J = 7.5 Hz), 7.37 (t, 2H, J = 7.6 Hz), 6.95 (d, 1H, J = 1.4 Hz), 6.85 (dd, 1H, J = 8.0, 1.4 Hz), 6.77 (d, 1H, J = 8.0 Hz), 6.63 (d, 1H, J = 15.8 Hz), 6.16 (dt, 1H, J = 15.8, 6.7 Hz), 5.96 (s, 2H), 4.86 (d, 2H, J = 6.7 Hz); 13C{1H} NMR (100 MHz, CDCl3): δ 153.8, 148.0, 147.8, 135.2, 132.9, 130.6, 128.5, 121.7, 120.2, 119.5, 108.3, 105.8, 101.1, 86.5, 80.5, 66.6; MS (ESI, pos) m/z (%): 329(6) [M + Na]+.

General Procedure—Pd-Catalyzed Decarboxylative Coupling: ()-((pent-1-en-4-yne-1,5-diylbis(benzene-4,1,2-triyl))tetrakis(oxy))tetrakis() (1′AA) [14]: A solution of 134 mg (0.171 mmol) of ester 2AA in 2.5 mL of freshly distilled THF was transferred under argon to a reaction tube containing 10 mg (0.0086 mmol) of Pd(P(Ph)3)4, and the tube was sealed. After heating for 4 h in an 80 °C oil bath, the contents of the tube were transferred to a round-bottomed flask with EtOAc, the solvent evaporated, and the residue subjected to flash chromatography (0–1% EtOAc/hexanes) to afford 1′AA as a yellow oil (105 mg, 0.142 mmol, 83%). 1H NMR and IR matched, which was previously reported [14]. 13C{1H} NMR (100 MHz, CDCl3): δ 147.3, 146.8, 146.6, 146.4, 130.9, 125.3, 124.3, 122.4, 121.0, 120.9, 119.6, 118.9, 116.5, 105.0, 85.2, 82.4, 26.0, 25.9, 22.9, 18.5, 18.4, −4.1. HRMS (ESI+) calculated for C41H71O4Si4 [M + H]+ 739.4424, found 739.4422.

()-((pent-1-en-4-yne-1,5-diylbis(4,1-phenylene))bis(oxy))bis() (1′BB): Prepared from 2BB following the general procedure described for 1′AA to afford 12 mg (22% yield) of a yellow oil. IR(ATR, neat): cm−1 2954, 2929, 2857, 2220, 1588, 1252, 1164, 906, 835, 779; 1H NMR (500 MHz, CD3Cl): δ 7.33–7.31 (m, 2H), 7.26–7.24 (m, 2H), 6.79–6.76 (m, 4H), 6.62 (d, 1H, J = 15.7 Hz), 6.12–6.07 (m, 1H), 3.32 (dd, 2H, J = 5.6, 1.6 H), 0.99 (s, 9H), 0.98 (s, 9H), 0.19 (s, 12H); 13C{1H} NMR (125 MHz, CDCl3): δ 155.5, 155.1, 132,9, 130.7, 130.5, 127.3, 122.4, 120.1, 120.0, 116.5, 85.5, 82.4, 25.7, 25.6, 22.9, 18.2, –4.4; MS (APCI, pos) m/z (%): 477(16) [M + H]+; HRMS (ESI+) calculated for C29H40O2Si2 [M + H]+ 477.2640, found 477.2640.

()-((pent-1-en-4-yne-1,5-diylbis(benzene-5,1,3-triyl))tetrakis(oxy))tetrakis() (1′CC): Prepared from 2CC following the general procedure described for 1′AA to afford 54 mg (56% yield) of a colorless oil. IR(ATR, neat): cm−1 2954, 2929, 2857, 1578, 1426, 1343, 1252, 1162, 1026, 827, 778; 1H NMR (500 MHz, CD3Cl): δ 6.57–6.50 (m, 3H), 6.50 (d, 2H, J = 2.2 Hz), 6.30 (t, 1H, J = 2.2 Hz), 6.23 (t, 1H, J = 2.2 Hz), 6.16 (dt, 1H, J = 15.7, 5.6 Hz), 3.32 (dd, 2H, J = 5.6, 1.7 Hz), 0.98 (s, 18H), 0.97 (s, 18H), 0.99 (s, 24H); 13C{1H} NMR (125 MHz, CDCl3): δ 156.6, 156.3, 138.9, 131.3, 124.6, 124.3, 116.8, 112.8, 111.6, 111.3, 86.2, 82,7, 25.7, 25.6, 22.9, 18.2, 18.1, −4.2, −4.4; MS (APCI, pos) m/z (%): 739(34) [M + H]+; HRMS (APCI, pos) calculated for C41H70O4Si4 [M + H]+ 739.4424, found 739.4396.

()-4,4’-(pent-1-en-4-yne-1,5-diyl)bis(1,2-difluorobenzene) (1DD): Prepared from 2DD following the general procedure described for 1′AA to afford 50 mg (28% yield) of 1DD as a yellow oil. IR(ATR, neat): cm−1 2930, 2204, 1599, 1513, 1430, 1290, 1216, 1114, 965, 872, 820, 773; 1H NMR (400 MHz, CDCl3): δ 7.25–7.06 (6H, m), 6.59 (1H, d, J = 15.7 Hz), 6.15 (1H, dtd, J = 15.7, 5.6), 3.34 (dd, 2H, J = 5.6, 1.3 Hz); 13C{1H} NMR (100 MHz, CDCl3): δ 150.4 (dd, J = 247.6, 12.9 Hz), 150.3 (dd, J = 250.6, 12.5 Hz), 149.9 (dd, J = 248.8, 12.9 Hz), 149.7 (dd, J = 248.8, 12.8 Hz), 134.2 (dd, J = 5.7, 4.1 Hz), 129.7, 128.2 (dd, J = 6.2, 3.6 Hz), 124.9 (d, J = 1.9 Hz), 122.4 (dd, J = 6.1, 3.4 Hz), 120.5 (d, J = 18.3 Hz), 120.3 (dd, J = 7.6, 4.2 Hz), 117.3 (d, J = 17.7 Hz), 117.2 (d, J = 17.3 Hz), 114.6 (d, J = 17.6 Hz), 86.9 (d, J = 1.3 Hz Hz), 81.0, 22.7; MS (ESI, pos) m/z (%): 289(8) [M − H]+; HRMS (ESI, pos) calculated for C17H10F4 [M]+ 290.0719, found 290.0714.

()-5,5’-(pent-1-en-4-yne-1,5-diyl)bis(benzo() (1EE): Prepared from 2EE following the general procedure described for 1′AA to afford 350 mg (80% yield) of 1EE as thin yellow needles. Mp = 58.2–60.0 °C; IR(ATR, neat): cm−1 2908, 2854, 1486, 1440, 1246, 1207, 1097, 1036, 967, 925, 813; 1H NMR (400 MHz, CD3COCD3): δ 7.03 (d, 1H, J = 1.7 Hz), 6.98 (dd, 1H, J = 8.0, 1.6 Hz), 6.91–6.90 (m, 1H), 6.88 (dd, 1H, J = 8.0, 1.7 Hz), 6.82 (dd, 1H, J = 8.0, 0.4 Hz), 6,79 (d, 1H, J = 8.0 Hz), 6.65 (dt, 1H, J = 15.7, 1.8 Hz), 6.18 (dt, 1H, J = 15.7, 5.7), 6.02 (s, 2H), 5.98 (s, 2H), 3.32 (dd, 2H J = 5.7, 1.8); 13C{1H} NMR (100 MHz, CD3COCD3): δ 149.0, 148.6, 148.4,148.0, 132.6, 131.6, 126.7, 123.5, 121.7, 117.9, 112.0, 109.1, 108.9, 106.3, 102.3, 102.0, 85.8, 82.2, 23.0; MS (APCI, pos) m/z (%): 307(29) [M + H]+; HRMS (CI+) calculated for C19H14O4 [M]+ 306.0892, found 306.0886.

()-6,6’-(pent-1-en-4-yne-1,5-diyl)bis(2,2-dimethyl-4) (1FF): Prepared from 2FF following the general procedure described for 1′AA to afford 4 mg (40% yield) of 1FF as a yellow oil: IR(ATR, CHCl3): cm−1 2988, 2923, 2220, 1613, 1582, 1496, 1384, 1265, 1202, 1142, 1117, 1065, 956; 1H NMR (500 MHz, CD3Cl): δ 7.24 (dd, 1H, J = 8.5, 1.9 Hz), 7.20 (dd, 1H, J = 8.5, 1.9 Hz), 7.08 (s, 1H), 6.99 (s, 1H), 6.76 (t, 2H, J = 9.0 Hz), 6.59 (d, 1H, J = 15.7 Hz), 6.08 (dt, 1H, J = 15.7, 5.7 Hz), 4.83 (s, 2H), 4.81 (s, 2H), 3.30 (dd, 2H, J = 5.7, 1.7 Hz), 1.54 (s, 12H); 13C{1H} NMR (125 MHz, CD3COCD3): δ 151.0, 150.7, 131.6, 130.7, 129.7, 128.0, 126.0, 122.5, 122.4, 119.4, 119.3, 117.2, 117.2, 115.5, 99.9, 99.6, 85.3, 82.4, 60.9, 60.6, 29.7, 24.7, 22.9; MS (APCI, pos) m/z (%): 391(1) [M + H]+, 423(5) [M + MeOH + H]+; HRMS (APCI, pos) calculated for C25H26O4 [M + H]+ 391.1904, found 391.1898.

()-((4-(5-(benzo()pent-4-en-1-yn-1-yl)-1,2-phenylene)bis(oxy))bis() (1′EA): Prepared from 2EA following the general procedure described for 1′AA to afford 52 mg (43% yield) of 1′EA as a yellow oil. IR(ATR, neat): cm−1 2929, 2857, 2193, 1593, 1557, 1505, 1488, 1407, 1308, 1250, 1099, 1038, 895, 828, 780; 1H NMR (500 MHz, CDCl3): δ 6.94–6.92 (m, 3H), 6.81 (dd, 1H, J = 8.0, 1.2 Hz), 6.76–6.74 (m, 2H), 6.60 (d, 1H, J = 15.7 Hz), 6.07 (dt, 1H, J = 15.7, 5.7 Hz), 5.95 (s, 2H), 3.31 (dd, 2H, J = 5.7, 1.5 Hz), 0.99 (s, 9H), 0.98 (s, 9H), 0.21 (s, 6H), 0.20 (s, 6H); 13C{1H} NMR (125 MHz, CDCl3): δ 147.9, 147.4, 146.9, 146.6, 131.7, 130.9, 125.3, 124.3, 122.8, 120.9, 120.7, 116.4, 108.2, 105.7, 101.0, 84.9, 82.6, 25.9, 22.9, 18.5, 18.4, −4.0, −4.1; (ESI, pos) m/z (%): 523(9) [M + H]+; HRMS (APCI, pos) calculated for C30H40O4Si2 [M + H]+ 521.2538, found 521.2532.

()-5-(5-phenylpent-1-en-4-yn-1-yl)benzo( (1EG) [32]: Prepared from 2EG following the general procedure described for 1′AA to afford 12 mg (20% yield) of 1EG as a yellow oil. IR(ATR, neat): cm−1 2916, 2895, 2212, 1502, 1488, 1444, 1279, 1248, 1167, 1124, 1099, 1035, 952, 927, 756; 1H NMR (400 MHz, CD3Cl): δ 7.46–7.44 (m, 2H), 7.31–7.29 (m, 3H), 6.93 (d, 1H, J = 1.5 Hz), 6.82 (dd, 1H, J = 8.0, 1.5 Hz), 6.75 (d, 1H, J = 8.0 Hz), 6.61 (dd, 1H, J = 15.6, 1.5 Hz), 6.08 (dt, 1H, J = 15.6, 5.7 Hz), 5.95 (2, 2H), 3.34 (dd, 2H, J = 5.7, 1.7 Hz); 13C{1H} NMR (125 MHz, CDCl3): δ 148.0, 147.0, 131.6, 131.0, 128.2, 127.8, 123.7, 122.5, 120.8, 108.2, 105.7, 101.0, 86.8, 82.8, 22.9; (ESI, pos) m/z (%): 263(3) [M + H]+.

General Procedure—Deprotection for Catechol-Containing Products: Rooperol (1AA) [14]: In a round-bottomed flask containing 44 mg of the silylester 1′AA (0.06 mmol) in 1 mL of dry DMF under Ar was placed 14 mg (0.24 mmol, 4 equivalent) of anhydrous KF. The flask was placed in an ice bath, and 0.2 mL of a solution prepared as a 1:100 dilution of 33% HBr/AcOH in DMF was added by syringe. After stirring for 1.5 h in the ice bath, the reaction mixture was diluted with EtOAc and washed with H2O and brine. The organic layer was dried over Na2SO4, filtered, and evaporated. The residue was subjected to flash chromatography 25% EtOAc/Hex + 1% MeOH) to afford 11 mg (65% yield) of catechol rooperol (1AA) as a yellow oil. 1H NMR (400 MHz, d-acetone): δ 6.93 (1H, brs), 6.89 (1H, d, J = 1.7 Hz), 6.80 (1H, dd, J = 8.0, 1.8 Hz), 6.74–6.77 (3H, m), 6.55 (1H, dt, J = 15.7, 1.7 Hz), 6.05 (1H, dt, J = 15.7, 5.7 Hz), 3.25 (2H, dd, J = 5.7, 1.8 Hz). 13C NMR (100 MHz, d6-acetone) δ 145.6, 145.0, 144.8, 130.9, 129.4, 123.7, 121.5, 118.3, 118.1, 155.2, 115.1, 114.9, 112.6, 84.1, 82.6, 22.1. IR (thin film) 3019, 1666, 1514, 1388, 1215, 755. MS (ESI-) m/z: 281 [M -H]–, 100); HRMS (ESI-) calculated for C17H13O4 [M − H]- 281.0819, found 281.0819.

()-4-(5-(benzo()pent-4-en-1-yn-1-yl)benzene-1,2-diol (1EA): Prepared from 1′EA following the general procedure described for 1AA to afford 2 mg (18% yield) of 1EA as a colorless oil. IR(ATR, neat): cm−1 2919, 2850, 2220, 1599, 1518, 1445, 1294, 1249, 1196, 1110, 1033, 962, 920, 817, 768; 1H NMR (500 MHz, CD3Cl): δ 6.96–6.92 (m, 3H), 6.82–6.79 (m, 2H), 6.75 (d, 1H, J = 8.0 Hz), 6.59 (dt, 1H, J = 15.6, 1.6 Hz), 6.06 (dt, 1H, J = 15.6, 5.7 Hz), 5.95 (s, 2H), 5.30 (s, 1H), 5.18 (s, 1H), 3.30 (dd, 2H, J = 5.7, 1.7 Hz); 13C{1H} NMR (125 MHz, CD3COCD3): δ 147.9, 147.0, 143.8, 143.0, 131.7, 130.9, 125.3, 122.7, 120.8, 118.6, 116.3, 115.3, 108.2, 105.6, 101.0, 85.1, 82.3, 22.9; (ESI, pos) m/z (%): 295(4) [M + H]+; HRMS (APCI, pos) calculated for C18H14O4 [M + H]+ 295.0965, found 295.0962.

General Procedure—Deprotection for Phenol- and Resorcinol-Containing Products: ()-4,4’-(pent-1-en-4-yne-1,5-diyl)diphenol (1BB) [33]: To a solution of 10 mg (0.019 mmol) of 1′BB in 0.2 mL of freshly distilled THF was added 4.7 µL (0.082 mmol) of AcOH. The solution was cooled in an ice bath, and 82 µL (0.082 mmol) of a 1M solution of TBAF in THF was added. After 20 min, EtOAc (5 mL) and water (5 mL) were added to the reaction mixture. The layers separated, and the aqueous layer was extracted 1 x EtOAc (5 mL). The combined organic layers were washed 1 x brine (5 mL). The combined aqueous layers were then back-extracted 1 x DCM (10 mL). The combined organics layers were dried over Na2SO4, filtered, evaporated, and subjected to flash chromatography 50% EtOAc/hexanes to afford 4 mg (71%) of 1BB as a colorless oil. IR(ATR, neat): cm−1 2955, 2851, 2224, 1602, 1506, 1238, 965, 831; 1H NMR (500 MHz, CD3Cl): δ 7.25–7.22 (m, 4H), 6.73–6.71 (m, 4H), 6.58 (d, 1H, J = 15.7 Hz), 6.07 (dt, 1H, J = 15.7, 5.7 Hz), 3.26 (dd, 2H, J = 5.7, 1.7 Hz); 13C{1H} NMR (125 MHz, CDCl3): δ 158.5, 158.0, 133.9, 131.9, 130.3, 128.4, 122.7, 116.3, 116.2, 116.0, 85.5, 83.6, 23.3; MS (ESI, neg) m/z (%): 249(100) [M − H]–.

()-5,5’-(pent-1-en-4-yne-1,5-diyl)bis(benzene-1,3-diol) (1CC): Prepared from 1′CC following the general procedure described for 1BB to afford 10 mg (42% yield) of 1CC as a colorless oil. IR(ATR, neat): cm−1 2960, 2925, 2853, 2220, 1587, 1503, 1441, 1340, 1299, 1142,997, 837; 1H NMR (500 MHz, CD3COCD3): δ 8.34 (s, 2H), 8.14 (s, 2H), 6.58 (dt, 1H, J = 15.7, 1.8 Hz), 6.44 (d, 2H, J = 2.2 Hz), 6.43 (d, 2H, J = 2.2 Hz), 6.35 (t, 1H, J = 2.2 Hz), 6.26 (t, 1H, J = 2.2 Hz), 6.20 (dt, 1H, J = 15.7, 5.6 Hz), 3.31 (dd, 2H, J = 5.6, 1.8 Hz); 13C{1H} NMR (125 MHz, CD3COCD3): δ 159.6, 159.4, 140.1, 132.2, 125.9, 125.0, 110.8, 105.7, 104.0, 102.8, 86.6, 83.7, 23.0; (ESI, pos) m/z (%): 283(6) [M + H]+; HRMS (APCI, pos) calculated for C17H14O4 [M + H]+ 283.0965, found 283.0965.

Cancer Cell Cytotoxicity Assays. HeLa, H460, and A549 cells were obtained from ATCC and grown in RPMI culture medium supplemented with 10% heat inactivated FBS (Life Technologies code 10270106). Cell culture media were supplemented with 4 mM glutamine (Lonza code BE17–605E), 100 μg/mL gentamicin (Lonza code 17-5182), and P/S (200 units/mL and 200 μg/mL) (Lonza code 17-602E) at 37 °C with 5% CO2. The effect of the investigated compounds on cell proliferation was determined by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. The compounds were dissolved in DMSO at a concentration of either 10 or 50 mM prior to cell treatment. The cells were trypsinized and seeded at various cell concentrations depending on the cell type. The cells were grown for 24–72 h, treated with test compounds at required concentrations, and incubated for 72 h in 100 or 200 μL media depending on the cell line used. Three replicates were performed. Cells treated with 0.1% DMSO were used as a negative control. The GI50 corresponds to the concentration of the compound of interest that reduces by 50% the growth of the cancer cell line of interest after having cultured it for 72 h in the presence of the compound in comparison to the untreated control condition.