Cholesterol → Camphene

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Total cholesterol, triglyceride and glucose levels in the serum were measured twice for each using a commercially available kit (Bio Clinical system), and insulin level was measured by ELISA using a mouse insulin kit (Shibayaki, Japan). Lipids were extracted from the liver tissue according to Folch et al.'s method. After adding 1 mL of distilled water to 0.25 g of liver tissue, the liver tissue was homogenized using a Polytron homogenizer (IKA-Werke GmbH & Co., Ultra-Turrax, Staufen, Germany). After adding 5 mL of chloroform:methanol solution (2:1, v/v) to the homogenate and mixing well, the mixture was centrifuged at 1000×g for 10 minutes. After adding 2 mL of chloroform:methanol solution (2:1, v/v) again to the supernatant, the same procedure was repeated to completely separate the lipid components of the liver. After adding 3 mL of chloroform:methanol:0.05% CaCl2 (3:48:47, v/v/v) solution to the remaining pellets and mixing well for 1 minute, followed by centrifugation at 1000×g for 10 minutes, the resulting pellets were completely dried with nitrogen gas. The dried lipids were dissolved in 1 mL of methanol and then analyzed. The same kit (Bio Clinical system) as the one used for the serum analysis was used to measure the triglyceride level of the liver tissue.

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