8-{[(E)-3-(2-Chlorophenyl)acryloyloxy]imino}-12,13-epoxytrichethec-9-en-4-yl (E)-3-(2-chlorophenyl)acrylate

In the title compound, C33H31Cl2NO6, the five-membered ring displays an envelope conformation, whereas the two six-membered rings both exhibit a chair conformation. As for the seven-membered ring, the dihedral angle between the mean planes formed by the four C atoms of the envelope unit and the three C and one O atoms of the six-membered chair is 69.08 (4)°, and these two mean planes are nearly perpendicular to the epoxy ring, making dihedral angles of 87.53 (4) and 88.67 (4)°, respectively.


Comment
The endophytic fungi Trichoderma taxi sp. nov. from Taxus mairei S. Y. Hu can produce a compound with fungicidal activity -Trichodermin (Zhang et al., 2007), which is a member of the 4β-acetoxy-12,13-epoxytrichothecene family (Nielsen et al., 2005). Bioassays showed Trichodermin strongly inhibited Rhizoctonia solani and Botrytis cinere. In order to find the relationship between the conjugated double bonds at 8 and 4 positions and biological activities, we designed to take the esterification reaction, thus, the title compound had been synthesized. Its molecular structure is shown in Fig. 1. In the molecule, the five membered ring displays an envelope conformation with atom C12 at the flap position 0.694 (5) Å out of the mean plane formed by C2, C3, C4 and C5. The pyranyl ring displays a chair conformation with the C11 and C12 atoms deviating by -0.578 (5) and 0.843 (5) Å from the mean plane formed by O1, C2, C5 and C6. It is interesting to note that the C8-C9-C10 bond angle is smaller than in the structure of Trichodermin (Chen et al., 2008) and Trichodermol (4α-hydroxy-12,13-epoxytrichothec-9-ene) (Cheng et al., 2009), two hydrogen bonds being displaced by the presence of the carbon-nitrogen double bond in the title compound. As for the seven-membered ring, the dihedral angle between the mean planes formed by C2-C3-C4-C5 and C2-C5-C6-O1 is 69.08 (4) °, two planes which are nearly perpendicular to the three-membered ring with angles of 87.53 (4) and 88.67 (4)°, respectively.

Experimental
In a flask, a solution of trichodermin (4.00 g, 13.70 mmoL) in 30 ml 1,4-dioxane was added dropwise over a period of 90 min to a refluxing solution of selenium dioxide (2.00 g, 18.02 mmoL) in 20 ml of 1,4-dioxane. After refluxing 12 h, the reaction was cooled and concentrated. Then 50 ml of ethyl acetate was added, and was washed with 5% aqueous sodium bicarbonate, dried, and concentrated in vacuum to 3.50 g of yellow liquid. The crude product was purified by flash column chromatography on silica gel using a mixture of petroleum ether and ethyl acetate (5: 1 by volume) as the eluent to give trichodermin-8-one (1.10 g, 26.2%) as colorless crystals. Then a mixture of trichodermin-8-one (1.00 g, 3.27 mmoL), hydroxylammonium chloride (0.45 g, 6.52 mmoL), and 30 ml methanol was stirred at 80 °C for 0.2 h. After the mixture was dissolved, a solution of potassium carbonate (4.00 g, 13.70 mmoL) in 20 ml water was added dropwise. The solution was stirred for 2.5 h, and concentrated, then extracted by ethyl acetate (10 ml) for three times, evaporated, and the crude product was purified via silica gel column chromatography using a 1: 2 (v/v) mixture of ethyl acetate and petroleum ether (boiling point range 60-90 °C) as the eluting solution to obtain (E)-trichodermin-8-hydroxy oxime as colorless crystals (0.89 g, 85.0%). At last, (E)-3-(2-chlorophenyl)acryloyl chloride (1.11 g, 5.54 mmoL) with 5 ml dichloromethane was added dropwise into a mixture of (E)-trichodermin-8-hydroxy oxime (0.89 g, 2.77 mmoL), triethylamine (33.8 mg, 0.28 mmoL), and N,N-dimethylpyridin-4-amine (0.56 g, 5.54 mmoL). The solution was stirred at room temperature and monitored by TLC. After 0.5 h, the mixture was washed with 1 N HCl, sat. NaHCO 3 and dried over anhydrous Na 2 SO 4 . The solvent was evaporated in vacuo to afford the crude product, which was purified by column chromatography to give the title compound (1.35 g, 80%) as a colorless solid. The solid was filtrated and recrystallized with acetone to get colourless blocks.

Refinement
The H atoms were geometrically placed and refined as riding, with C-H = 0.93 Å and U iso (H) = 1.2U eq (C) for aromatic H atoms, with C-H = 0.97 Å and U iso (H) = 1.2U eq (C) for methylene H atoms, with C-H = 0.98 Å and U iso (H) = 1.2U eq (C) for methine H atoms, and with C-H = 0.96 Å and U iso (H) = 1.5U eq (C) for methyl H atoms. Fig. 1. The molecular structure of (I) showing the atom-labelling scheme. Displacement ellipsoids are drawn at the 40% probability level.

Special details
Experimental Refinement. Refinement of F 2 against ALL reflections. The weighted R-factor wR and goodness of fit S are based on F 2 , conventional R-factors R are based on F, with F set to zero for negative F 2 . The threshold expression of F 2 > σ(F 2 ) is used only for calculating Rfactors(gt) etc. and is not relevant to the choice of reflections for refinement. R-factors based on F 2 are statistically about twice as large as those based on F, and R-factors based on ALL data will be even larger.

Fractional atomic coordinates and isotropic or equivalent isotropic displacement parameters (Å 2 )
x y z U iso */U eq