2. Simulated salivary electrolyte stock solution: Transfer 15.1 mL
of 0.625 M KCl, 3.7 mL of 0.625 M KH 2 PO 4 , 6.8 mL of
1.25 M NaHCO 3 , 0.5 mL of 0.187 M MgCl 2 6H 2 O, and
0.06 mL of 0.625 M (NH 4 ) 2 CO 3 to a 500 mL graduated
cylinder with 100 mL water. Make up to 500 mL and mix.
Store at À20
C.
3. Simulated gastric electrolyte stock solution: Transfer 6.9 mL of
0.625 M KCl, 0.9 mL of 0.625 M KH 2 PO 4 , 12.5 mL of
1.25 M NaHCO 3 , 11.8 mL 2.5 M NaCl, 0.4 mL of 0.187 M
MgCl 2 6H 2 O, and 0.5 mL of 0.625 M (NH 4 ) 2 CO 3 to a
500 mL graduated cylinder with 100 mL water. Make up to
500 mL and mix. Store at À20
C.
4. Simulated intestinal electrolyte stock solution: Transfer 6.8 mL
of 0.625 M KCl, 0.8 mL of 0.625 M KH 2 PO 4 , 42.5 mL of
1.25 M NaHCO 3 , 9.6 mL 2.5 M NaCl, and 1.1 mL of
0.187 M MgCl 2 6H 2 O to a 500 mL graduated cylinder with
100 mL water. Make up to 500 mL and mix. Store at À20
C.
5. Stock solution of 1500 U of α-amylase per mL of simulated
salivary electrolyte solution (see Notes 5 and 6).
6. Stock solution of 25,000 U of porcine pepsin per mL of
simulated gastric electrolyte stock solution (see Notes 5 and 7).
7. Stock solution of 800 U of pancreatin from porcine pancreas
per mL of simulated intestinal electrolyte stock solution (see
Notes 5 and 8).
8. Stock solution of 0.16 mmol of porcine bile extract per mL of
simulated intestinal electrolyte stock solution (see Notes 9 and
10).
9. Laboratory blender with glass container (1–2 L capacity) with
speed control and timer.
10. 100 mL GL45 clean glass laboratory bottles.
11. Magnetic stirrer with 4–6 stirring positions. Cross type PTFE
magnetic stir bars 25 mm diameter.
12. Incubator chamber with adjustable temperature control to
0.1
C and internal power sockets for stirring unit(s).
2.3 Measurement
and Adjustment of pH
during In Vitro
Digestion
1. Adjustment of pH values at the initial and during the gastric
and intestinal stages is made with 1 M HCl or 1 M NaOH.
2. The pH measurements should be performed with a pH electrode suitable for difficult samples containing colloids, emulsions, creams, or very viscous samples (see Note 11).
2.4 Assessment of
Lipolysis Efficiency
1. Folch solution, chloroform–methanol 2:1 (v/v), is made fresh
each time.
2. 1 M HCl, 1 M NaCl, and H 2 O–H 2 SO 4 (1:1; v/v).
Carotenoid In Vitro Digestion
379
of 0.625 M KCl, 3.7 mL of 0.625 M KH 2 PO 4 , 6.8 mL of
1.25 M NaHCO 3 , 0.5 mL of 0.187 M MgCl 2 6H 2 O, and
0.06 mL of 0.625 M (NH 4 ) 2 CO 3 to a 500 mL graduated
cylinder with 100 mL water. Make up to 500 mL and mix.
Store at À20
C.
3. Simulated gastric electrolyte stock solution: Transfer 6.9 mL of
0.625 M KCl, 0.9 mL of 0.625 M KH 2 PO 4 , 12.5 mL of
1.25 M NaHCO 3 , 11.8 mL 2.5 M NaCl, 0.4 mL of 0.187 M
MgCl 2 6H 2 O, and 0.5 mL of 0.625 M (NH 4 ) 2 CO 3 to a
500 mL graduated cylinder with 100 mL water. Make up to
500 mL and mix. Store at À20
C.
4. Simulated intestinal electrolyte stock solution: Transfer 6.8 mL
of 0.625 M KCl, 0.8 mL of 0.625 M KH 2 PO 4 , 42.5 mL of
1.25 M NaHCO 3 , 9.6 mL 2.5 M NaCl, and 1.1 mL of
0.187 M MgCl 2 6H 2 O to a 500 mL graduated cylinder with
100 mL water. Make up to 500 mL and mix. Store at À20
C.
5. Stock solution of 1500 U of α-amylase per mL of simulated
salivary electrolyte solution (see Notes 5 and 6).
6. Stock solution of 25,000 U of porcine pepsin per mL of
simulated gastric electrolyte stock solution (see Notes 5 and 7).
7. Stock solution of 800 U of pancreatin from porcine pancreas
per mL of simulated intestinal electrolyte stock solution (see
Notes 5 and 8).
8. Stock solution of 0.16 mmol of porcine bile extract per mL of
simulated intestinal electrolyte stock solution (see Notes 9 and
10).
9. Laboratory blender with glass container (1–2 L capacity) with
speed control and timer.
10. 100 mL GL45 clean glass laboratory bottles.
11. Magnetic stirrer with 4–6 stirring positions. Cross type PTFE
magnetic stir bars 25 mm diameter.
12. Incubator chamber with adjustable temperature control to
0.1
C and internal power sockets for stirring unit(s).
2.3 Measurement
and Adjustment of pH
during In Vitro
Digestion
1. Adjustment of pH values at the initial and during the gastric
and intestinal stages is made with 1 M HCl or 1 M NaOH.
2. The pH measurements should be performed with a pH electrode suitable for difficult samples containing colloids, emulsions, creams, or very viscous samples (see Note 11).
2.4 Assessment of
Lipolysis Efficiency
1. Folch solution, chloroform–methanol 2:1 (v/v), is made fresh
each time.
2. 1 M HCl, 1 M NaCl, and H 2 O–H 2 SO 4 (1:1; v/v).
Carotenoid In Vitro Digestion
379