Three New Triterpene Esters from Pumpkin (Cucurbita maxima) Seeds

Three new multiflorane-type triterpene esters, i.e. 7α-hydroxymultiflor-8-ene-3α,29-diol 3-acetate-29-benzoate (1), 7α-methoxymultiflor-8-ene-3α,29-diol 3,29-dibenzoate (2), and 7β-methoxymultiflor-8-ene-3α,29-diol 3,29-dibenzoate (3), were isolated from seeds of Cucurbita maxima, along with the known compound, multiflora-7,9(11)-diene-3α,29-diol 3,29-dibenzoate (4). Compound 1 exhibited melanogenesis inhibitory activities comparable with those of arbutin. In cytotoxicity assays, compounds 1 and 3 exhibited weak cytotoxicity, with IC50 values of 34.5–93.7 μM against HL-60 and P388 cells.

Cucurbita maxima (English name: squash, pumpkin, Japanese name: Kabocha) is indigenous to the plateaus of central and south America, but is nowadays cultivated throughout the world. Its fruits,

Plant Material
The seeds of Cucurbita maxima, produced in India, were purchased from Takada Seeds Co., Ltd. (Osaka, Japan) in 2011. A voucher specimen was deposited in the Herbarium of the Laboratory of Medicinal Chemistry, Osaka University of Pharmaceutical Sciences.

Cell Cultures
The cell lines HL-60 (human leukemia) and P388 (murine leukemia) were grown in RPMI 1640 medium, while B16 4A5 cells were grown in D-MEM. The medium was supplemented with 10% FBS and antibiotics (100 units/mL penicillin and 100 μg/mL streptomycin). The cells were incubated at 37 °C in a 5% CO 2 humidified incubator.

Determination of B16 4A5 Cells Proliferation
The assay of B16 4A5 cells proliferation was examined according to a method reported previously [9].

Assay of Melanin Content
The assay of melanin content was performed as described previously [9].

Cytotoxicity Assay against Cancer Cell Lines
The cytotoxicity assay was determined previously [19].