A method for evaluating and verifying biochemical methane potential test completion performed with landfilled municipal solid waste.

The biochemical methane potential (BMP) test is significant for the landfill industry as it provides a means to evaluate the gas potential, and therefore potential degradability, of both incoming and in-place municipal solid waste (MSW). However, the BMP test is not standardized making comparison of BMP results across sites problematic. For example, the BMP test duration has historically ranged from 20 days to several months with most current BMP tests lasting 60 days. However, the gas generation data can potentially be modelled for any of those durations to produce a prediction of the ultimate BMP value (BMPULT). Currently, the predicted BMPULT values of 23 long-duration (115-150 days) BMP tests were used to determine the required quantity of data (i.e. number of days) needed to produce an accurate BMPULT prediction. Results showed that no single test duration produced both accurate and efficient results, so a novel performance-based endpoint was proposed. The relative change in predicted BMPULT values with respect to time (dBMPULT/dt) was chosen as a potential performance-based completion metric. Results indicate that once the absolute normalized dBMPULT/dt value is within <2.5, <1.5 and <0.6% day-1 that the predicted BMPULT is within 20, 10 and 5% of the true BMPULT, respectively. Overall, the use of performance-based metrics for determining BMP test completion will allow for the collection of partial data sets, reduced experimental times and verification of results.

Binding of active Ras and its mutants to the Ras binding domain of PI-3-kinase: A quantitative approach to KD measurements.

Ras family GTPases (H/K/N-Ras) modulate numerous effectors, including the lipid kinase PI3K (phosphatidylinositol-3-kinase) that generates growth signal lipid PIP3 (phosphatidylinositol-3,4,5-triphosphate). Active GTP-Ras binds PI3K with high affinity, thereby stimulating PIP3 production. We hypothesize the affinity of this binding interaction could be significantly increased or decreased by Ras mutations at PI3K contact positions, with clinical implications since some Ras mutations at PI3K contact positions are disease-linked. To enable tests of this hypothesis, we have developed an approach combining UV spectral deconvolution, HPLC, and microscale thermophoresis to quantify the KD for binding. The approach measures the total Ras concentration, the fraction of Ras in the active state, and the affinity of active Ras binding to its docking site on PI3K Ras binding domain (RBD) in solution. The approach is illustrated by KD measurements for the binding of active H-Ras and representative mutants, each loaded with GTP or GMPPNP, to PI3Kγ RBD. The findings demonstrate that quantitation of the Ras activation state increases the precision of KD measurements, while also revealing that Ras mutations can increase (Q25L), decrease (D38E, Y40C), or have no effect (G13R) on PI3K binding affinity. Significant Ras affinity changes are predicted to alter PI3K regulation and PIP3 growth signals.