*3.1. Identification of Anthocyanins by HPLC*/*MS*

The WBB powder used to prepare the products contained at least 22 anthocyanins (Table 1), which were identified by comparing their mass-to-charge (*m*/*z*) values and elution orders with

previous studies [4,5,34]. Blueberries are unique in that three different sugars (galactose, glucose, arabinose) are commonly attached to the five anthocyanidins (delphinidin, cyanidin, petunidin, peonidin, malvidin) [34]. This was confirmed in our study; however, we were unable to detect peonidin-3-arabinoside using our HPLC method. We were unable to obtain complete separation of all of the anthocyanins present in the extract due to the complexity of the anthocyanin profile. Peak 15 contained two co-eluting compounds, namely cyanidin-3-(6′′-malonyl) galactoside and cyanidin-3-(6′′-acetyl) galactoside, and peak 18 was composed of three co-eluting compounds, namely delphinidin-3-rutinoside, cyanidin-3-(6′′-malonyl) glucoside, and malvidin-3-(6′′-acetyl) galactoside. We were unable to identify peak 17, which appeared to be a delphinidin derivative based on its aglycone *m*/*z* of 303, but the molecular ion *m*/*z* value was ambiguous. Many of the anthocyanins were present in acylated form. Two of the cyanidin glycosides (galactoside and glucoside) were acylated with malonic acid, whereas delphinidin, cyanidin, and malvidin galactosides as well as petunidin, peonidin, and malvidin glucosides were acylated with acetic acid moieties.


**Table 1.** Peak assignments, retention times (RT in min), and mass spectral data of anthocyanins in extract from WBB powder.
