Monte Carlo sampling can be used to determine the size and shape of the steady-state flux space

Constraint-based modeling results in a convex polytope that defines a solution space containing all possible steady-state flux distributions. The properties of this polytope have been studied extensively using linear programming to find the optimal flux distribution under various optimality conditions and convex analysis to define its extreme pathways (edges) and elementary modes. The work presented herein further studies the steady-state flux space by defining its hyper-volume. In low dimensions (i.e. for small sample networks), exact volume calculation algorithms were used. However, due to the #P-hard nature of the vertex enumeration and volume calculation problem in high dimensions, random Monte Carlo sampling was used to characterize the relative size of the solution space of the human red blood cell metabolic network. Distributions of the steady-state flux levels for each reaction in the metabolic network were generated to show the range of flux values for each reaction in the polytope. These results give insight into the shape of the high-dimensional solution space. The value of measuring uptake and secretion rates in shrinking the steady-state flux solution space is illustrated through singular value decomposition of the randomly sampled points. The V(max) of various reactions in the network are varied to determine the sensitivity of the solution space to the maximum capacity constraints. The methods developed in this study are suitable for testing the implication of additional constraints on a metabolic network system and can be used to explore the effects of single nucleotide polymorphisms (SNPs) on network capabilities.