Increase in Plasma Concentrations of Geranylgeranoic Acid after Turmeric Tablet Intake by Healthy Volunteers

Geranylgeranoic acid (GGA) is one of the most potent cancer-preventive acyclic retinoids. GGA has been shown to induce cell death in human hepatoma-derived HuH-7 cells. We have recently reported the natural occurrence of GGA and its related compounds in several medicinal herbs such as turmeric, basil, rosehip, cinnamon and others [Shidoji and Ogawa, J. Lipid Res., 45: 1092–1103, 2004]. In the present study, we performed oral administration of turmeric tablets to healthy volunteers in order to investigate bioavailability of natural GGA. By using liquid chromatography/mass spectrometry, authentic GGA was eluted at a retention time of around 18 min as a negative ion of m/z 303.4. With healthy volunteers, plasma GGA was detected prior to the tablet intake and its concentrations were increased at 2 h after its intake and maintained at higher level until 4 h, suggesting an efficient bioavailability of preformed GGA in the turmeric tablets through oral administration. These results indicated that GGA in the turmeric tablet was absorbed as an intact form from intestinal mucosa. The present study provides a clue to conduct a research for cancer preventive roles of GGA in a number of spices

Polished rice as natural sources of cancer-preventing geranylgeranoic acid

Geranylgeranoic acid, a 20-carbon polyprenoic acid (all-trans 3,7,11,15-tetramethyl-2,4,6,10,14-hexadecatetraenoic acid) and its derivatives were previously developed as synthetic “acyclic retinoids” for cancer chemoprevention. Recently, we demonstrated the natural occurrence of geranylgeranoic acid in various medicinal herbs (Shidoji and Ogawa, 2004). In this present study, we present several lines of evidence to demonstrate that geranylgeranyl diphosphate taken in foods could be metabolized to GGA through geranylgeraniol and geranylgeranyl aldehyde via the following steps: 1) The conversion from geranylgeranyl diphosphate to geranylgeraniol was demonstrated to occur by the action of bovine intestinal alkaline phosphatase, with a Km of 46.1 µM. 2) Geranylgeraniol oxidase-mediated conversion of geranylgeraniol to geranylgeranyl aldehyde was revealed in rat liver homogenates, which activity was mainly localized in the mitochondrial fraction. The mitochondrial enzyme showed a Km of 92.9 µM. 3) The conversion of geranylgeranyl aldehyde to geranylgeranoic acid by geranylgeranyl aldehyde dehydrogenase in rat liver homogenates was absolutely dependent on exogenously added NAD+ or NADP+. The Km of the mitochondrial geranylgeranyl aldehyde dehydrogenase was 27.5 µM for geranylgeranyl aldehyde. Taken together, our data suggest that cancer preventive geranylgeranoic acid could be a physiological metabolite from commonly consumed foods

Imprinting analysis by droplet digital PCR coupled with locked nucleic acid TaqMan probes

Imprinted genes are differentially expressed in a parent-of-origin-specific manner. Parental origin of the alleles is discriminated by intragenic DNA polymorphisms. Comparisons of parental allelic expression have been analysed by semiquantitative RT-PCR. Here, we developed a novel quantitative method for allelic expression of the imprinted gene Ube3a, which inactivation and mutations cause Angelman syndrome and predominantly expressed by the maternal allele in neuronal tissues. In this method, cDNA was amplified by droplet digital PCR (ddPCR) coupled with allele-specific locked nucleic acid (LNA) TaqMan probes, which labelled by FAM and HEX were designed to detect the SNPs in the target regions. ddPCR assay demonstrated that the sense transcript of Ube3a was equally expressed from both parental alleles in adult tissues except neuronal tissues, where Ube3a expression from the paternal allele was about 10 to 14% of total Ube3a expression in adult brain, and 20% in spinal cord. The antisense transcript of Ube3a was expressed at 60% to 70% of the sense transcript of Ube3a in adult brain. Changes in the Ube3a transcripts during postnatal brain development were also evaluated by ddPCR. The ddPCR method is far more reliable and simpler to use than semiquantitative PCR to analyse skewed or faint allelic expression of imprinted genes