*2.4. In Vitro Digestion*

The microfluidic noodle was cut into < 2 mm pieces before being subjected to the simulated in vitro digestion. The protocol for digestion was referred from the INFOGEST method with minor modification [32]. All simulated saliva fluid (SSF), simulated gastric fluid (SGF) and simulated intestinal fluid (SIF) solutions were prepared according to the INFOGEST method. Specifically, SSF was comprised of 15.1 mmol/L potassium chloride (KCL), 3.7 mmol/L potassium dihydrogen phosphate (KH2PO4), 13.6 mmol/L sodium bicarbonate (NaHCO3), 0.15 mmol/L magnesium chloride hexahydrate (MgCl2(H2O)6) and 0.06 mmol/L ammonium carbonate ((NH4)2CO3); SGF was comprised of 6.9 mmol/L KCL, 0.9 mmol/L KH2PO4, 25 mmol/L NaHCO3, 47.2 mmol/L NaCl, 0.1 mmol/L MgCl2(H2O)6 and 0.5 mmol/L (NH4)2CO3; SIF was comprised of 6.8 mmol/L KCL, 0.8 mmol/L KH2PO4, 85 mmol/L NaHCO3, 38.4 mmol/L NaCl and 0.33 mmol/L MgCl2(H2O)6. To start, 5 g of cut noodle samples were mixed with 3.5 mL SSF, 0.5 mL salivary a-amylase (1500 U/mL) solution, 25 uL CaCl2 (0.3 M) and 975 uL deionized water, and were incubated in the water bath at 37 ◦C, 85 rpm for 2 min. Gastric digestion was started with the oral digesta being acidified, by adding 1 M hydrogen chloride (HCL) (~0.2 mL) to adjust the pH to reach 3.0. It was subsequently mixed with 7.5 mL SGF, 5 uL CaCl2 (0.3 M), 1.6 mL pepsin

(25,000 U/mL) solution and 695 uL deionized water, and was incubated in the water bath at 37 ◦C, 85 rpm for 1.5 h. Afterwards, to simulate the intestinal digestion, 1 M sodium hydroxide (NaOH) (~0.15 mL) was used to adjust the pH to 7.0, and then was mixed with 11 mL SIF, 5 mL pancreatin solution (800 U/mL), 2.5 mL bile (10 mM), 40 uL CaCl2 (0.3 M) and 1.31 mL deionized water, and was incubated in the water bath at 37 ◦C, 85 rpm for 2.5 h. Right after the final stage of the simulated gastrointestinal digestion was completed, raw digesta was centrifuged at 19,802× *g*, 20 ◦C for 45 min. Given that large particles are unable to pass through the mucus layer and be taken up by epithelium cells [33], the aqueous supernatant after filtration (pore size 0.45 μm) was assumed to be comprised of mixed micelles where the lutein solubilized [34]. We considered the lutein solubilized in the micellar fractions as the bioaccessible lutein. During the entire digestion procedure, all the samples were kept in the amber color tubes or the containers were covered with aluminum foil to minimize the photodecomposition of lutein.

#### *2.5. Extraction and Quantification of Lutein*

Lutein in digesta, micelle fraction and homogenate were extracted and analyzed as previously reported [35]. Briefly, digesta, micellar fraction or homogenate was extracted with acetone:petroleum ether (1:1, *<sup>v</sup>*/*<sup>v</sup>*, second and third times was extracted with petroleum ether alone), vortexed for 2 min and was centrifuged for 10 min at 19,802× *g*, 20 ◦C. The supernatant layer was collected and the above extraction was repeated three times. All the supernatant layers were combined, and then it was evaporated by nitrogen gas. The final samples were reconstituted in methanol:methyl tert-butyl ether (MTBE) (1:1, *v*/*v*) and were filtered through a 0.45 μm filter. The extraction procedure was fully carried out under dull red light, and 0.1% butylated hydroxytoluene ( *w*/*v*) was added in the extraction solvents to minimize lutein degradation.

Lutein was detected by the HPLC (Waters, US) at 4 ◦C at the wavelength of 450 nm with a YMC carotenoid C30 column, 250 mm × 4.6 mm ID (YMC, Japan), that has been reported previously [35]. The mobile phases were comprised of methanol:MTBE:water (A, 81:15:4, *v*/*v*/*v*) and methanol:MTBE:water (B, 9:87:4, *v*/*v*/*v*). The gradient program was carried out as follows: an initial condition of eluent A:B was 100:0 (%), then there was a linear increase till A:B was 81:19 (%) at 3 min, followed by an A:B of 47:53 (%) at 25 min, and then a rapid increase till A:B was 0:100 (%) at 27 min, held for 10 min and finally back to the initial condition in 3 min, allowing for a 10 min hold as re-equilibration. The flow rate was set as 1 mL/min and the injection volume was 80 μL.
