3.3.4. Juice

The total anthocyanin content of the juice decreased with storage time for each storage temperature (*p* < 0.0001). The total anthocyanin content of juice stored at 4.4 ◦C and 21 ◦C is shown in Figure 2. After eight weeks of storage, the juice stored at 4.4 ◦C retained 90.7% of total anthocyanins compared with control samples (Day 0), whereas the juice stored at 21 ◦C retained 69.1%. Concentrations of anthocyanins are known to readily decline during storage of blueberry juice at ambient temperature [23,36], but refrigeration is an effective treatment to ameliorate anthocyanin losses [29,37,38]. Changes in the major individual anthocyanins in the juice stored at 4.4 ◦C and 21 ◦C over eight weeks of storage are shown in Figure S4. At 4.4 ◦C, peonidin-3-galactoside, cyanidin-3-arabinoside, malvidin-3-galactoside, malvidin-3-glucoside, and malvidin-3-(6′′-acetyl) galactoside remained stable over the eight weeks of storage. At 4.4 ◦C, all anthocyanins showed >50% retention, with the minimal percent retention being 57.7% for the unknown delphinidin derivative. This compound, however, did not significantly decrease over storage at 21 ◦C, along with the two co-eluting anthocyanins (cyanidin-3-(6′′-malonyl) galactoside + cyanidin-3-(6′′-acetyl) galactoside). Besides these two compounds, the percent retention of anthocyanins at 21 ◦C ranged from 59% (malvidin-3-(6′′-acetyl) glucoside) to 75.5% (petunidin-3-glucoside).

#### 3.3.5. Gummy Product

The total anthocyanin content of the gummy product decreased with storage time for each storage temperature (*p* < 0.0001). The total anthocyanin content of the gummy product stored at 4.4 ◦C and 21 ◦C is shown in Figure 2. After eight weeks of storage, the gummy product stored at 4.4 ◦C and 21 ◦C retained 43.2% and 50.6%, respectively, of their original total anthocyanin content (Day 0). Consistent with our findings, levels of total anthocyanins declined in gelatin gels prepared with grape pomace extract over 24 weeks of storage at 21 ◦C, with losses most pronounced in gels exposed to neon light [39]. Maier et al. (2009) [39] also reported similar retention of total anthocyanins in gels stored for 24 weeks at 6 ◦C and 24 ◦C. The lower amount of anthocyanins recovered in the gummies stored at 4.4 ◦C compared to the same product stored at 21 ◦C may be explained by reduced extraction efficiency due to the hardening of the gel at low temperature, as opposed to degradation late during storage. Changes in the major individual anthocyanins in the gummy product stored at 4.4 ◦C and 21 ◦C over eight weeks of storage are shown in Figure S5. At 4.4 ◦C, all the individual anthocyanins decreased with storage time with retentions <50%, except for the two co-eluting anthocyanins (cyanidin-3-(6′′-malonyl) galactoside + cyanidin-3-(6′′-acetyl) galactoside) (60.9%) and malvidin-3-glucoside (52.1%). The percent retentions of the rest of the anthocyanins at this storage temperature ranged from 29.3% to 49.2%. When stored at 21 ◦C, two anthocyanins did not significantly decrease with time, namely the unknown delphinidin derivative and the two co-eluting compounds (cyanidin-3-(6′′-malonyl) galactoside + cyanidin-3-(6′′-acetyl) galactoside). For the rest of the anthocyanins, percent retentions ranged from 40% (cyanidin-3-glucoside and cyanidin-3-galactoside) to 71% (malvidin-3-(6′′-acetyl) glucoside).

In all the products, the individual anthocyanin loss did not appear to be impacted by the anthocyanidin structure or the type of sugar moiety attached (data not shown).

#### 3.3.6. Product Comparison of Anthocyanin Composition over Eight Weeks of Storage

The distribution of the products according to their individual anthocyanin profile as affected by storage time (0, 2, 4, 6, and 8 weeks) can be visualized on a PCA scores plot (Figure 3). The first principal component (PC1) explained 83.9% of the variation with all the individual anthocyanins being positively loaded on PC1. Therefore, PC1 represents the amount of individual anthocyanins. The juice and ice pop samples had high scores on PC1 (except for the juice samples stored at 21 ◦C for 6 and 8 weeks). The oatmeal bar samples also had positive scores on PC1 for the earlier storage times, whereas the oatmeal bar samples stored at 21 ◦C for eight weeks were the only oatmeal sample to have

a negative score. Except for the control samples (Day 0), all the graham cracker cookie samples had negative scores on PC1, regardless of the storage temperature. Finally, all gummy samples (for both storage at 4.4 ◦C and 21 ◦C) had negative scores on PC1, with scores becoming smaller with storage time. The PCA figure confirmed higher values of anthocyanins in the juice and ice pop samples, as well as, to a lesser extent, the oatmeal bars. The graham cracker cookie and gummy samples did not demonstrate high values for anthocyanins, with a clear loss of anthocyanins with storage time for the gummy samples.

− **Figure 3.** Principal component analysis (PCA) scores plot of blueberry products stored at −20 ◦C (ice pop), 4.4 ◦C and 21 ◦C (graham cracker cookie, gummy, juice, oatmeal bar) for 0, 2, 4, 6, and 8 weeks (*n* = 3/time point). −

#### *3.4. Changes in Percent Polymeric Color in Blueberry Products during Storage*

Changes in percent polymeric color (% PC) values in oatmeal bar, graham cracker cookie, juice, and gummy product stored at 4.4 ◦C and 21 ◦C over eight weeks of storage are shown in Figure 4.

**Figure 4.** Stability of percent polymeric color in blueberry stored at 21 ◦C and 4.4 ◦C (*n* = 3/time point). Shaded area around lines represents 95% confidence intervals for predicted values.

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