Optimizing fabrication parameters via Taguchi method for production of high yield hydroxyapatite microsphere scaffolds using Drop-on-Demand inkjet method

Abstract

Drop on demand (DOD) inkjet method is a cost-efficient way of producing hydroxyapatite (HAp) microsphere scaffolds with narrow size distribution. However, DOD fabrication parameters may influence the yield and characteristics of the microsphere scaffolds. Testing different permutations and combinations of fabrication parameters is costly and time consuming. Taguchi method could be used as a predictive tool for optimizing the key fabrication parameters to produce HAp microspheres with desired yield and properties, minimizing the number of experimental combinations to be tested. The aim of this study is to investigate the influence of the fabrication parameters on the characteristics of the microspheres formed and determine optimum parameter conditions for producing high yield HAp microsphere scaffolds with the desired properties intended to serve as potential bone substitutes. We aimed to achieve microspheres with high production yield, microsphere size of <230 μm, micropore sizes <1 μm, rough surface morphology and high sphericity. Experiments were conducted using Taguchi method with a L9 orthogonal array at three levels per parameter to determine optimum parameter values for (1) operating pressure, (2) shutter speed duration, (3) nozzle height and (4) CaCl₂ concentration. Based on signal-to-noise (S/N) ratio analysis, the identified optimum parameter conditions for operating pressure, shutter speed duration, nozzle height and CaCl₂ concentration to be 0.9-1.3 bar, 100 ms, 8 cm and 0.4 M respectively. The microspheres obtained had an average size of 213 μm, 0.45 μm micropore size, high sphericity index of 0.95 and high production yield of 98%. Confirmation tests and ANOVA results affirms the validity of Taguchi method in optimizing HAp microspheres with high yield, desired size, micropore size and shape. HAp microsphere scaffolds produced by optimum conditions were subjected to a 7-day in-vitro study. Cells remained viable and continued to proliferate (increased 1.2-fold) over 7 days with microspheres maintaining high cell density with cells bridging between microspheres. Alkaline phosphatase (ALP) assay increased 1.5-fold from day 1, suggesting good osteogenic potency of HAp microspheres as potential bone substitutes.

Keywords: Taguchi method; bone tissue engineering; cell culture; hydroxyapatite; microspheres; optimization parameters.