2. Materials and methods2.1. Sample collection and preparationRJ composition varies according to its origin (Messia, Caboni, &Marconi, 2005). In this study, there were three types of RJ: 1) eightfreshly harvested RJ samples obtained from eight apiaries in ZhejiangProvince in June 2010, when naturally available forage wascomplemented by protein (bee pollen) and sugar syrup; 2) fourfresh RJ samples harvested from Hubei Province during the floweringperiod of Camellia sinensis in April 2011; and 3) four fresh RJsamples harvested from Qinghai Province during the floweringperiod of rape (Brassica campestris) in July 2011. All samples wereproduced by Italian bees (Apis mellifera ligustica) and stored at <4 Cpost-harvest. Following homogenization, each sample was transferredto 40 air-tight glass bottles. These bottles were randomlyassigned to four groups with 10 bottles per group; each group wasstored at different temperatures: 18 C, 4 C, 20 C ± 2 C (roomtemperature or RT), and 30 C. Samples were collected at differenttime points: 0, 7, 14, 21, 28, 35, 42, 57, and 98 days of storage. Eachbottle was sampled only once, to avoid the potential effect ofrepeated freeze-thaw deterioration in the sample.To verify whether the method was reliable for the determinationof RJ freshness, eight RJ samples were obtained from a majorChinese bee-product company in Zhejiang Province, who storestheir samples at 18 C for no longer than 36 h post-harvest.Additionally, eight commercial RJ samples were obtained fromsupermarkets in Beijing.
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