Avanti j e

The pulsed electric field-treated bacterial suspension and nontreated bacteria were diluted in BHI medium at a ratio of 1:10 and incubated at 37 °C for 6 h. The cultures were collected and centrifuged at 4000× g for 10 min at 4 °C (Avanti J-E centrifuge, Beckman Coulter, Atlanta, CA, USA). The sediment was washed three times with 1 mL PBS and the sediment of the fourth centrifuge was stored at −80 °C.

Non-acidified samples were centrifuged at 10,000 rpm for 10 min using a Beckman Coulter Avanti J-E series centrifuge unit (Beckman Coulter, Brea, CA, USA). Collected pellets were re-suspended in approximately 10 mL of supernatant and stored at −80 °C. DNA from pellet biomass was extracted using the MP Bio FastDNA Extraction Kit for Soil (MP Biomedicals, Solon, OH, USA) as per the manufacturer’s instructions. Extracted gDNA was quantified using fluorometry (Qubit 2.0 fluorometer; sample volume 1 μL). Sample gDNA concentrations were not normalized to preserve possible less abundant taxa.

Three replicates of approximately six liters of milk from Alpine goats were collected from the bulk tank of the milking parlor at the International Goat Research Center at Prairie View A&M University. Fresh goat milk was brought to the laboratory and divided into two batches of WM and SM. The goat skim milk was prepared by centrifugation (Avanti J-E centrifuge, Beckman Coulter Inc., Indianapolis, IN, USA) using 100 mL centrifuge tubes at 6200× g at 4 °C for 10 min. The fucoxanthin with 20% purity was dissolved into HPLC grade ethanol and mixed with both goat raw whole and skim milk to a final concentration of 10.67 μg/mL (2.56 mg/240 mL of milk, which is one serving). Samples of WM and SM without fucoxanthin were used as the controls. Both raw WM and SM were pasteurized by LTLT (low temperature—long time) at 64 °C for 30 min (Safgard Pres-Vac Home Pasteurizer Model P-3000; the Schlueter Co., Janesville, WI, USA). The samples of whole or skim milk with and without fucoxanthin were stored under refrigeration temperature (4 °C) for storage stability studies. All samples that were used for the analyses of milk composition, quantification of fucoxanthin, determination of pH and titratable acidity, evaluation of color, or the measurement of lipid oxidation were pasteurized by LTLT. A flow chart of goat milk processing and physicochemical analyses is given in Figure 1.

Dried plant powders were used for extraction with two different solvents, including ethanol and distilled water. Each dried powder sample was suspended in the solvent at a ratio of 1:10 (w/v) and kept in an ultrasonic incubator (S100H, Elma GmbH, Singen, Germany) at 45 °C for 30 min. The mixture was swirled at 200 rpm for 60 min at room temperature using an orbital incubator shaker (Gyromax™ 747 Incubator Shaker, Amerex Instruments, Concord, CA, USA). The supernatant was separated from the plant residues by centrifugation (Avanti J-E, Beckman Coulter, Brea, CA, USA) at 10,000 rpm for 20 min at 10 °C. The supernatant was then filtered through Whatman No. 1 filter paper (Whatman, Buckinghamshire, UK). The filtrate was entirely dried in a vacuum rotary evaporator (HS-2005S-N, Hahnshin, Gimpo-si, Korea) or lyophilized (UniFreeze FD-8, Daihan Scientific, Wonju, Korea), depending on whether the solvent was ethanol or water, respectively. The dried extracts were stored at 4 °C. The crude extracts were dissolved in dimethyl sulfoxide (DMSO) to obtain stock solutions of 100 mg/mL. These stock solutions were then diluted to different concentrations for further analysis.

Fresh cultures were obtained after activation by three successive transfers in MRS broth. Cultures in late‐log phase (10¹⁰ cfu/ml) were harvested by centrifugation at 3,000 × g for 10 min (Avanti J‐E, Beckman Coulter), washed with a sterile peptone solution (0.1%, w/v). The final wet cell mass was weighed and dispersed in peptone solution to obtain suspension with 10¹⁰ cfu/ml cells.

Fresh garlic bulbs were peeled, washed, chopped, and homogenized with distilled water at a ratio of 1:2 (garlic:distilled water) and then stirred for 12 h at room temperature before subjected to filter through cheesecloth and centrifuged at 6000g (Avanti® J-E; Beckman Coulter, TX). The clear supernatant was collected and freeze-dried. The commercial pickled garlic was washed with tap water twice to remove the excess pickled solution and then drained for 2 min before brought to peel, wash, chop, homogenize and extract as mentioned in fresh garlic extract.