Lomatia tasmanica, W. M. Curtis

Deans, Bianca J., Tedone, Laura, Bissember, Alex C. & Smith, Jason A., 2018, Phytochemical profile of the rare, ancient clone Lomatia tasmanica and comparison to other endemic Tasmanian species L. tinctoria and L. polymorpha, Phytochemistry 153, pp. 74-78 : 76-77

4.3. Extraction of Lomatia tasmanica

Diethyl ether maceration. Fresh leaves including petiole of L. tasmanica (17.6 g) (extracted within 1 h of obtaining plant material to reduce the likelihood of decomposition) were cut at the petiole into ∼ 2.5 cm pieces (to form an even layer of material in the extraction vessel). The plant material was then completely submerged in diethyl ether (250 mL) and the vessel was sealed and maintained at room temperature. After 5 h, the leaves were observed to change to a black color, while the solution displayed a vibrant orange color. The mixture was filtered, dried (Na 2 SO 4), filtered, and evaporated under reduced pressure to provide an orange residue (193 mg). This mixture was purified by automated flash chromatography (0 30 → % hexanes/EtOAc) to deliver juglone (60 mg, 0.34% w/w) as red/orange crystalline solid (NMR spectroscopic data is provided in the Supporting Information) and a colorless semi-solid (53 mg, 0.30% w/w) containing a mixture of nonacosane (major component) and heptacosane (minor component).

Nonacosane. ( Lytovchenko et al., 2009). EI-MS (m / z): 408. Heptacosane. ( Lytovchenko et al., 2009). EI-MS (m / z): 380. See Supporting Information for 1 H NMR spectrum and the GC chromatogram of the mixture containing nonacosane and heptacosane.

PHWE. Immediately following the above-mentioned diethyl ether maceration, residual solvent was allowed to evaporate from the leaf material that remained (∼10 min). The ensuing L. tasmanica leaves (14.1 g) were then coarsely ground in a spice grinder, mixed with sand (2 g), placed into the portafilter (sample compartment) of an espresso machine and extracted using 35% v/v EtOH/H 2 O (200 mL of a hot solution). The ensuing extract was then concentrated under reduced pressure on a rotary evaporator to dryness (50 ̊C bath temperature) to provide a brown residue (1.20 g). The crude extract was adsorbed onto a ∼1:1 mixture of silica gel and Celite and fractionated by automated flash column chromatography (0 → 40% MeOH/EtOAc; 18 min). This resulted in a combined fraction predominantly containing the two saccharides (539 mg) as judged by 1 H and 13 C NMR spectroscopy. Due to difficulties isolating these compounds by flash column chromatography, this mixture was reacted with acetic anhydride to enable purification as described immediately below.

Peracetylation of saccharides obtained from PHWE. A portion of the mixture (206 mg), obtained as described immediately above, was dissolved in pyridine (1.5 mL), cooled to 0 ̊C. Acetic anhydride (1.5 mL) was added dropwise and the reaction mixture magnetically stirred at room temperature. After 21 h, the reaction mixture was concentrated under reduced pressure to afford a brown gum (440 mg). Purification by automated flash column chromatography (0 → 100% EtOAc/hexanes; 10 min) provided an inseparable mixture of α- and β- glucose pentaacetate (85 mg) as a colorless film (NMR spectroscopic data is provided in the Supporting Information).