Calcium chloride

A calcination solution of urea (Carl Roth GmbH) and calcium chloride of various concentrations (Carl Roth GmbH) was prepared for MICP. The cell suspension of S. pasteurii was harvested after cultivation to an optical density of 2.0 and stored sterile at 4°C. Quartz sand from Haltern, Germany with a quartz content of 98 % was used for this study. Particle sizes ranged from 125 μm to 500 μm with the largest percentage of particles (43%) having a size between 125 and 250 μm. For consolidation of silica sand, cell suspension was first added by the printer to the sand through a nozzle with a diameter of 200 μm under a pressure of 0.69 bar. For the negative control NH4‐YE medium was added instead of cell suspension under the same conditions. Then, after 15 min, an equivalent volume of calcination solution was added. This was followed by a period of 3.5 h during which the calcination solution used was allowed to react. Before each cycle the solution was washed with deionized water to wash off excess salts from the previous cycle. Between the intervals the container was closed with a layer of plastic film to prevent the sample from drying out. Following the sixth printing interval, the sand bed was oven dried for 48 h at 55°C. Subsequently, the sand bed was treated with Resazurin Red S for staining the calcium carbonate.

Native and PLGA-loaded BNC were investigated for their fluid-releasing capabilities using a method previously reported by Zahel et al. [42 (link)]. Briefly, an aqueous solution containing 142 mmol L⁻¹ sodium chloride as well as 2.5 mmol L⁻¹ calcium chloride (both Carl Roth) was prepared. Subsequently, 35% (w/w) gelatin (Bloom 180, Carl Roth) was suspended in this solution and incubated at 60 °C until fully dissolved. Afterward, Petri dishes with a diameter of 100 mm were filled with 37 g of the gelatin solution each, tightly sealed, and incubated at 25 °C for 3 h to allow the gelation of gelatin. Subsequently, the Petri dishes were weighed, the BNC samples were placed on the gelatin surface afterward and the dishes were tightly sealed. After 48 h of incubation, the BNC samples were removed, and the dishes were weighed again. The fluid release (FR) was calculated according to Equation (5):
where m_after represents the weight of the Petri dishes after incubation, m_before the weight of the Petri dishes before incubation, and A the contact area between BNC fleece and gelatin gel. Experiments were performed in triplicates and repeated once.