Targeted Extraction of Select Natural Products Using Supercritical Carbon Dioxide
Creators: name: Kramp, Kari
Publisher: name: Université d'Ottawa / University of Ottawa
Year: 2010
Subjects: subject: Biology, Molecular.
subject: Chemistry, Organic.

description: This thesis examined the supercritical carbon dioxide (CO2) extraction (SFE) of targeted secondary metabolites from selected natural products. Phytochemical characterization and determination of biological efficacy was completed using advanced analytical methods and in-vivo bio-assays. More than 500 SFE extracts were generated and characterized. Sample preparation (particle size, moisture content) and extraction parameters (pressure, temperature and flow rate) were investigated and the ability of SFE to extract specific biosynthetic classes of compounds was evaluated. Where warranted, pilot scale studies were conducted for larger scale biological trials. SFE of the neotropical vine, Souroubea sympetala, a traditional medicine used for a culture bound syndrome related to anxiety, resulted in an extract rich in betulinic acid (5.54 +/- 0.24 mg/g) determined by HPLC-APCI/MS. When subjected to validated rodent anxiety behavioral assays (e.g. elevated plus maze (EPM)) the SFE extract demonstrated significant anxiolysis (anti-anxiety) compared to the vehicle control, with a 50% increase in time spent in open arms and a 73% increase in unprotected head dips observed. The significant in-vivo anxiolysis observed provides scientific support for the ethnobotanical use of this traditional medicine and a promising lead for a natural health product (NHP) to treat anxiety. Systematic variation of SFE parameters (pressure and temperature) on pyrethrum (Chrysanthemum cinerariifolium) oleoresin, a natural insecticide, resulted in a residue free pyrethrin concentrate, suitable for organic agricultural applications. The highest pyrethrin concentration (0.531 g/g) and highest ratio of pyrethrins I (pyrethrin I, cinerin I, jasmolin I) to pyrethrins II (pyrethrin II, cinerin II, jasmolin I) (PI:PII = 1.95) was obtained at 40 °C; 10 MPa while 40°C; 30 MPa produced the lowest pyrethrin concentration (0.436 g/g) and lowest ratio of PI:PII (1.87). Total pyrethrin recovery was highest at 40°C; 30 MPa (8.39 g pyrethrins) vs. 40°C; 10 MPa (6.96 g pyrethrins). The insecticidal activity of the SFE extracts of the pyrethrum oleoresin was confirmed (&gt;70% mortality at 48 h) using the Colorado potato beetle (CPB) a major pest of agriculture. SFE of piperamides from black pepper (Piper nigrum), a promising botanical insecticide and synergist, was successful at pressures above 30 MPa and temperatures greater than 50&amp;dot;C with total yields greater than 7.87% (CV &lt; 0.10) and piperamide concentrations greater than 150 mg/g (CV &lt; 6.0). Flow rate, sample size, and particle size were optimized at 5 L/min, 10 g, and 1 mm mesh respectively. Insecticidal activity corroborated dose dependent piperamide bioactivity. Extract from 10 MPa; 60°C (42.38 mg/g) produced 10.0+/-6.8 (24 h) and 13.3+/-6.7% (48 h) mortality. Extract from 35 MPa; 40°C (148.56 mg/g) produced 70.0+/-10.0 (24 h) and 73.3+/-8.4% (48 h) mortality. Northern shrimp (Pandelus borealis) by-products extracted by SFE (35 MPa; 40°C) generated a deep red oil rich in polyunsaturated fatty acids (PUFAs), specifically eicosapentaenoic acid EPA (7.8+/-0.06%) and docosahexaenoic acid DHA (8.0+/-0.07%). Optimization of pressure (60 MPa), temperature (80°C), moisture (13%), particle size (0.85 mm), and flow rate (9 L/min) resulted in a more efficient extraction without significantly changing concentrations of EPA and DHA. Pilot scale studies confirmed experimental data and supports further investigation of commercial production of an omega -3 (o-3) concentrate from Northern shrimp by-products.