1. William Kim Science Research – Period 4 Mrs. Flanagan

2. Figure 1: Number of people age 65 and over, by age group, selected years 1900-2006 and projected 2010-2050 Need

3. Figure 2: Life expectancy at ages 65 and 85, by sex, selected years 1900-2004 Need

4. Knowledge Base http://www.mayday-info.dk/715.0.html

5. Knowledge Base http://www.imsc.res.in/~sitabhra/research/ neural/celegans/celegans.gif http://shirleywho.files.wordpress.com/2008 /09/c-elegans_esa.jpg

6. Fig. 3 Blueberry polyphenols extend lifespan and slow aging in Caenorhabditis elegans. Literature Review Wilson et. al. 2006

7. Literature Review  James Joseph (2007-2009)  Tested and still testing fruit polyphenols and their effects on aging and the brain  Tested effects of blackberries, blueberries, strawberries, plum juice, and other berry fruits

8. Literature Review  life cycle of C. elegans is comprised of the embryonic stage, four larval stages (L1-L4) and adulthood  The end of each larval stage is marked with a molt where a new, stage-specific cuticle is synthesized and the old one is shed (Cassada and Russell, 1975) http://www.sfu.ca/biology/faculty/ hutter/hutterlab/research/pictures /Ce_life_cycle.gif

9. Purpose Therefore the purpose of this experiment is to test the ability of blueberries, blackberries, and other fruit polyphenols in the prolonging of lifespan in C. elegans. Hypothesis Null Hypothesis: The blueberries, etc. will have no effect in prolonging the lifespan of C. elegans. Alternate Hypothesis: The blueberries, etc. will have a significant effect in prolonging the lifespan of C. elegans.

10. Polyphenols and Aging Methodology Caenorhabditis elegans Short lifespan of about 2-3 weeks, rapid generation time, and experiemental flexibilty Control Medium Sample Size = 5 Medium + Fed crude blueberry extract (Vaccinium angustifolium) Sample Size = 5 Measurement Average pumping rate of pharynx contraction is recorded with the use of Motic Plus Camera attached to a stereomicroscope. Statistical Analysis Statistical analysis by T-test, p<.05, will be used to determine if the collected data is significant. Medium + Fed crude blackberry extract Sample Size = 5 Medium + Fed crude strawberry extract Sample Size = 5 Medium + Fed crude Plum extract Sample Size = 5

11. Protocols for Measurement  Commercially prepared single strength wild blueberry juice (Vaccinium angustifolium) is applied to a preconditioned C18 Column  C18 column is washed with water to remove fructose, glucose, and organic acids of blueberries, then with 100% methanol to obtain the total polyphenolic fraction http://www.waters.com/waters/partDetail.htm?local e=101&partNumber=WAT066224 http://www.abnativeplants.com/index.cfm/fu seaction/plants.plantdetail/plant_ID/17/index.htm Low Bush Blueberries

12. Protocols for Measurement  Fruit polyphenol extracts is added to NGM agar plates of C. elegans on the first day of the lifespan assay  Lifespan scoring is initiated after hermaphrodites complete their final larval molt, on the first day of adulthood  Pharynx pumping rates are measured on days 4, 6, 8, and 10 of C. elegans adulthood

13. Protocols for Measurement  Pharynx contraction rate recorded with Motic Plus camera attached to a stereomicroscope  Pumps per minute measured http://www.technicalvideo.com/ photos/ElegansPharynx.jpg

14. Doability  Motic Plus with Stereomicroscope available in lab for pharynx contraction observation  Fruit polyphenols are easily obtainable  C. elegans are easily obtainable and culturable

15. BudgetVendor Catalogue # PageItem Item # Quantit y Unit $$ Total $$ Carolina Science 2009 78248 C. elegans N2 (wild type) NP-17- 3500 1 $10.95 (per culture) $10.95 Caroline Science 2009 78248 Nematode Growth Agar NP-17- 3520 5 $6.25 (per 135 ml bottle) $31.25 https://www.wat ers.com/waters/ partDetail.htm?p artNumber=WA T066224 n/an/a Symmetry C18 Column, 3.5 µm, 4.6 x 75 mm WAT066 224 1$543.00$543.00 Carolina Science Websiten/an/aMethanol 8749501 $5.75 per 500 mL $5.75 Total Price: $590.95

16. Bibliography “Aging Statistics.” Administration on Aging. (2009) 24 March 2009. “Antioxidants.” McGraw-Hill Yearbook of Science & Technology. McGraw-Hill, 2001. “Antioxidants.” MedlinePlus. (2009) 9 Feb. 2009. Carlson, Rick E., Kirby, Brett S., Voyles, Wyatt F., and Dinenno, Frank A. “Evidence for impaired skeletal muscle contraction-induced rapid vasodilation in aging humans.” American Journal of Physiology – Heart and Circulatory Physiology 294 (2008): H1963-H1970. Joseph, James. "ARS Project: The effects of walnuts on age related changes in neuroprotection, inflammatory signaling and behavior (414597).". Kenney, Richard A. Physiology of Aging: A Synopsis. Chicago, London. Year Book Medical Publishers, INC, 1982. Larsson, L., Grimby, G., and Karlsson, J.. “Muscle Strength and Speed of Movement in Relation to Age and Muscle Morphology.” Journal of Applied Physiology 46 (1979): 451-456. "NCLD - Dyspraxia." NCLD - Home. 04 May 2009. Oliver, Rush H., Albury, Acchia N.J., and Mousseau, Timothy A.. “Programmed Cell Death in Flight Muscle Histolysis of the House Cricket.” Journal of Insect Physiology 53 (2007): 30-39. Scalbert, Augustin, Johnson, Ian T., and Saltmarsh, Mike. “Polyphenols: antioxidants and beyond.” The American Journal of Clinical Nutrition 81 (2005): 215S- 7S. “Types of contractions.” Muscle Physiology. (2006). University of California. 31 May 2006. USDA/Agricultural Research Service. "Adding Walnuts To Good Diet May Help Older People Improve Motor And Behavioral Skills." ScienceDaily 25 April 2009. 5 May 2009. “What Is an Antioxidant?.” American Dietetic Association. (2006). 14 Sept. 2006.. Wilson, Mark A., Shukitt-Hale, Barbara, Kalt, Wilhemina, Ingram, Donald K., Joseph, James A., and Wolkow, Catherine A. “Blueberry polyphenols increase lifespan and thermotolerance in Caenorhabditis elegans.” Aging Cell. 5 (2006): 59-68.