Source Paper
Elucidating the Mechanism of the Liqi Yangyin Formula in Treating Depression–Constipation Comorbidity: An Integrative Approach Using Network Pharmacology and Experimental Validation
Xu L, Ong SS, Deng X, Qian Y, Tang Z et al.
Pharmaceuticals (Basel) • 2026
First Black Stool Test
Objective: To measure intestinal transit time by assessing the time from charcoal administration to first black fecal pellet appearance
Protocol Steps
Prepare charcoal suspending vehicle
Dissolve gum acacia in distilled water with heating until clear, add kaolin, boil three times, cool to room temperature, and adjust to final volume
View evidence from paper
“A charcoal solution was prepared by dissolving gum acacia (1 g) in 80 mL distilled water with heating until clear, adding kaolin (5 g), boiling the mixture three times, cooling to room temperature, and adjusting to 100 mL with distilled water”
Prepare activated carbon suspension
Prepare activated carbon suspension using the suspending vehicle and mix thoroughly before use
View evidence from paper
“An activated carbon (charcoal) suspension (e.g., 5% w/v) was prepared with this vehicle and mixed thoroughly before use”
Fast mice
Fast mice for 24 hours while allowing free access to water
View evidence from paper
“After a 24 h fast (water ad libitum), mice received the charcoal solution”
Administer charcoal solution
Give charcoal solution by oral gavage at a dose based on body weight and record administration time
View evidence from paper
“mice received the charcoal solution by oral gavage at 0.1 mL per 10 g body weight. The administration time (T1) was recorded”
House individually and observe
House animals individually for observation and monitor for first black fecal pellet
View evidence from paper
“Animals were then housed individually for observation. The administration time (T1) and time of first black fecal pellet (T2) were recorded”
Calculate time to first black stool
Calculate the time difference between administration and first black stool appearance
View evidence from paper
“Time (min) = T2 − T1”