Solubility Analysis

4.3.1. Laboratory Stage Mixer

Solutions of MPC (5% w/w) were made by placing an octagonal magnetic stir bar (8 x 38mm) in a 250 ml beaker, weighing 95.0 g DI H2O in the beaker, placing 5.0 g MPC in a weigh dish, transferring the MPC to the DI H2O, and hand-mixing (100 clockwise revolutions with a thin spatula), prior to reconstitution using a laboratory stage mixer (R010 Power, IKA Works, Wilmington, NC). Each was allowed to reconstitute at 960 rpm (speed setting #8) at 23 °C ± 1.0 °C for 1 h and 3 h. One complete block was tested at a time, and two replicates of each treatment were tested at a time. Aliquots (13 ml) of these samples were transferred to a series of 15 ml falcon tubes and centrifuged at 700 x g for 10 min at 23 °C. The supernatant was separated from the pelleted material by withdrawing supernatant into a pipette. Percent solubility was calculated as supernatant total solids (TS) divided by bulk solution TS, multiplied by 100. TS was determined using CEM LabWave 9000 (CEM Corp, Matthews, NC) using parameters of 100%

power and 4 min drying time, after running duplicate samples under vacuum oven drying (AOAC 927.05) to verify accuracy of LabWave 9000 measurements. Measurements were conducted in duplicate. A diagram of this experiment is shown in

Figure 13.

Figure 13: Diagram of laboratory stage mixer solubility experiment

4.3.2. Insolubility Index

One-hundred ml of water (24 ˚C ± 0.2 ˚C) was poured into the mixing jar (bottom diameter = 7.11 cm, blade diameter = 5.08 cm) (Waring Commercial Blender [Model 34B197, 120 Volts AC, 50 Hz to 60 Hz, 7.0 Amps], Torrington, CT). An aliquot of MPC (5 g) was placed into the mixing jar. The analysis then proceeded according to Insolubility Index: GEA Niro analytical method A 3 a (Anonymous, 2010c). Three drops of Antifoam B Emulsion (Sigma-Aldrich, St. Louis, MO) were added, and mixing commenced at 3800 rpm for 90 sec. The solution was allowed to sit for 15 min. After 15 min. elapsed, the solution was stirred with a thin spatula and transferred to two 50 ml centrifuge tubes. Centrifugation took place at 910 rpm for 5 min. A pipette was used to dispose of all sediment-free liquid more than 5 ml above the sediment layer. The centrifuge tubes were then filled with DI water to the 50 ml mark, the sediment was dispersed with a thin spatula, and centrifugation again took place at 910 rpm for 5 min.

Centrifugation (700 x g)

Labwave 9000

Stage mixer and 8 beakers Transfer 13 ml to falcon tubes Centrifugation Obtain TS of supernatant Beaker

95g DI H2O 5g MPC

Stir (100 x clockwise revolutions

Either 1 hour or 3 hours total mixing time

Obtain TS of bulk solution

The amount of sediment remaining was reported in ml. Measurements were conducted in duplicate.

4.3.3. Solubility Using Modified Method of Anema et al. (2006)

Solutions of MPC (5% w/w) were made in a 400 ml beaker by weighing 95.0 g DI H2O in the beaker, placing 5.0 g MPC in a weigh dish, transferring the MPC to the DI H2O, and hand-mixing (100 clockwise revolutions with a thin spatula). This solution was placed into a 30 °C water bath. Mixing immediately commenced at 600 rpm (speed setting #5) using a propeller-blade (blade diameter = 4.4 cm) attached to an overhead stirrer (StedFast Stirrer [Model SL600], Fisher Scientific, Tustin, CA). After 30 min., samples of MPC solutions were then withdrawn, and the supernatant was obtained by centrifugation at 700 x g for 10 min. Percent solubility was calculated as supernatant total solids TS divided by bulk solution TS, multiplied by 100. TS was determined using CEM LabWave 9000 (CEM Corp, Matthews, NC) using parameters of 100% power and 4 min.

drying time. Measurements were conducted in duplicate.

4.3.4. Solubility in pH-adjusted Environments

Solutions of MPC (5% w/w) were made by placing an octagonal magnetic stir bar (8 x 38mm) in a 250 ml beaker, weighing 95.0 g DI H2O in the beaker, placing 5.0 g MPC in a weigh dish, transferring the MPC to the DI H2O, and hand-mixing (100 clockwise revolutions with a thin spatula), prior to reconstitution using a laboratory stage mixer (R010 Power, IKA Works, Wilmington, NC). Each was allowed to reconstitute at 960 rpm for 10 min. to allow for dispersion of material. pH of all samples were recorded.

through 3 to pH of treatment level 4. Samples were allowed to mix at 23 °C ± 1.0 °C until 3 h total mixing time was achieved. Samples were checked at 30 min. intervals to ensure consistency in pH, and readjusted with 1% NaOH if necessary. Aliquots (13 ml) of these samples were transferred to a series of falcon tubes and centrifuged at 700 x g for 10 min at 23 °C. The supernatant was separated from the pelleted material by withdrawing supernatant into a pipette and transferring to a 15 ml falcon tube. Percent solubility was calculated as supernatant TS divided by bulk solution TS, multiplied by 100. TS was determined using CEM LabWave 9000 (CEM Corp, Matthews, NC) using parameters of 100% power and 4 min. drying time. Measurements were conducted in duplicate. Figure 14 diagrams this experiment.

In a separate experiment, solutions of MPC (5% w/w) were made by placing an octagonal magnetic stir bar (8 x 38mm) in a 250 ml beaker, weighing 95.0 g DI H2O in the beaker, placing 5.0 g MPC in a weigh dish, transferring the MPC to the DI H2O, and hand-mixing (100 clockwise revolutions with a thin spatula), prior to reconstitution using a laboratory stage mixer (R010 Power, IKA Works, Wilmington, NC). Each was allowed to reconstitute at 960 rpm for 10 min. to allow for dispersion of material. pH of all samples were recorded. While mixing, 1% HCl solution was used to adjust pH of samples treatment levels 2 through 4 to pH of treatment level 1. Samples were allowed to mix at 23 °C ± 1.0 °C until 3 h total mixing time was achieved. Samples were checked at 30 min. intervals to ensure consistency in pH, and readjusted with 1% HCl if necessary.

Aliquots (13 ml) of these samples were transferred to a series of falcon tubes and centrifuged at 700 x g for 10 min at 23 °C. The supernatant was separated from the pelleted material by withdrawing supernatant into a pipette. Percent solubility was

calculated as supernatant total solids TS divided by bulk solution TS, multiplied by 100.

TS was determined using CEM LabWave 9000 (CEM Corp, Matthews, NC) using parameters of 100% power and 4 min drying time. Measurements were conducted in duplicate. Figure 14 diagrams this experiment.

Figure 14: Solubility of experiment testing solubility in pH-adjusted environments