University of Pennsylvania Department of Bioengineering Objectives & Hypotheses: Analysis of the Effect of Calcium Content on Chicken Bone Fracture Energy - Group WB5 Crucial Role of Calcium in Biological Systems Hydroxyapatite: Primary Mineral Component in Bone [Ca 5 (PO 4 ) 3 OH] Objective 1: Examine the Effects of Calcium Content on Fracture Strength in Chicken Bones Objective 2: Examine the Precision Between Two Analytical Methods Expanding on Experiment 4: dimensional  compositional Calcium Removed and Content Quantified Via Spectrophotometry and Gravimetric Analysis. Hypothesis 1: Calcium Content Will Significantly Correlate to Fracture Strength in Chicken Bones in Each Method. (p<0.05; Via Regression Analysis) Hypothesis 2: Both Methods Will Provide a Statistically Equivalent Finding on Correlation of Calcium Content to Fracture Strength (p<0.05; Via Two- Tailed t-Test)

University of Pennsylvania Department of Bioengineering Methods, Protocol & Equipment: Fracture 8 chicken leg specimens using the pendulum to determine the fracture energy (see lab 4). Use saw to create two 1cm rings from each broken half and measure their combined mass and volume for each specimen. Use MicroMill Grinder on each pair of rings. Combine 1 gram of ground bone from each specimen into 8 beakers. Pour 50mL of RDF into beakers and let sit for one hour. Use CaCl 2(s) and H 2 O, mixed with 13mL EDTA (aq) and 1mL of Eriochrome Black T indicator to make ten samples with volumes of 50mL. Pipette 5mL into cuvettes and create a calibration curve. Transfer 5ml of each solution and 1ml of indicator to 8 cuvettes and test on spectrophotometer for absorbance. Supersaturate remaining solutions with 200mL of Na 2 CO 3(aq), to precipitate CaCO 3(s) to find mass and moles of Ca after weighing.

University of Pennsylvania Department of Bioengineering Proposed Results/Findings/Deliverables: Potential Pitfalls: Chicken bone fracture energy table Calcium content of bone analysis table Statistical Analysis:-Is there a calcium to E fracture correlation? -Are the two methods of calcium density analysis consistent? A limited range of calcium density data (limits statistical power) Spectrophotometer and mass measurements disagree Unanticipated chemical interactions

University of Pennsylvania Department of Bioengineering Materials and Budget & Justification: New Equipment MicroMill Grinder- BEL1576 (Scienceware) 1, Rapid Decalcification Formulation (RDF), 1 gal (Statlab) $47.00 Calcium Chloride, 200g- C Eriochrome Black T, 100g-E G (Sigma) Ethylenediaminetetraacetic acid (EDTA), 1kg (Sigma) Sodium Carbonate, 1kg- S7795 (Sigma) Total $ Major Equipment Pendulum Spectrophotometer Other Lab Equipment Beakers, cuvettes