Antidepressant-like effects of the hydroalcoholic extracts of Hemerocallis citrina and its potential active components

Abstract

Background: Herbal therapies are potential alternatives and adjuncts for depression treatment. The present study aims to investigate the antidepressant-like effects of hydroalcoholic Hemerocallis citrina extracts and its potential neuropharmacological components.

Methods: Hydroalcoholic H. citrina extracts were phytochemically analyzed. Behavioral models, including tail suspension tests and open field tests, were performed to evaluate the antidepressant-like effects of the extracts. A possible mechanism was explored by analyzing brain monoamine neurotransmitters. Toxicity and histopathological analyses were performed to determine whether or not the extracts are safe for oral administration.

Results: The antidepressant-like effects of hydroalcoholic H. citrina extracts were mainly related to flavonoids, especially rutin and hesperidin. The extract prepared using 75% ethanol (i.e., HCE75) exhibited the highest active flavonoid content and activity. Orally administered 400 mg/kg of HCE75 significantly induced an antidepressant-like effect, whereas the combination of equivalent rutin and hesperidin dosages exhibited the same profiles. Isobologram analysis showed sub-additive antidepressant interactions between rutin and hesperidin. HCE75 (400 mg/kg, p.o.) increased the serotonin and dopamine levels in the central nervous system. Mortality and lesions were not observed upon oral administration of up to 5000 mg/kg HCE75.

Conclusions: The antidepressant-like effects of hydroalcoholic H. citrina extracts are mainly related to flavonoids, especially rutin and hesperidin. The serotonergic and dopaminergic systems may have major roles. The active extract is toxicologically safe for oral administration.

Figure 1

Flavonoid fingerprint of HCE75 as a representative HPLC chromatogram. The peaks correspond to (1) rutin, (2) hesperidin, (3) quercitrin, and (4) quercetin.

Figure 2

Effects of HCE on immobility time in TST (A) and line crossings in OFT (B). The apparent correlations between (C) chemical and (D) flavonoid constituent dosages and the immobility time in HCE in the TST. The results are expressed as% immobility time relative to the control group (vehicle) for the antidepressant-like effects. The mice were tested 60 min after they were administered (p.o.) with the vehicle (physiological saline with 2% Tween 80), fluoxetine (20 mg/kg), and HCE (400 mg/kg). Data are expressed as mean ± S.E.M. (n = 10). Data were analyzed using one-way ANOVA for multiple comparisons, followed by post hoc Student–Newman–Keuls test. *p < 0.05, **p < 0.01, and ***p < 0.001 compared with the control group (vehicle); and #p < 0.05, ##p < 0.01, and ###p < 0.001 compared with the positive reference group (fluoxetine). Linear regression was performed along with the ordinary least squares estimation.

Figure 3

Effects of the pure rutin (A), hesperidin (B), quercetin (C), and quercitrin (D) on immobility time in TST. The mice were tested 60 min after they were administered (p.o.) with the vehicle (physiological saline with 2% Tween 80), 20 mg/kg fluoxetine, 0.1, 1, 2, 4, and 8 mg/kg rutin, 0.03, 0.3, 1, 2, and 4 mg/kg hesperidin, 0.01, 0.1, 0.2, 0.4, and 0.8 mg/kg quercetin, and 0.01, 0.1, 0.2, 0.4, and 0.8 mg/kg quercitrin. Data are expressed as mean ± S.E.M. (n = 10). Data were analyzed using one-way ANOVA for multiple comparisons, followed by post hoc Student–Newman–Keuls test. *p < 0.05, **p < 0.01, and ***p < 0.001 compared with the control group (vehicle); #p < 0.05, ##p < 0.01, and ###p < 0.001 compared with the positive reference group (fluoxetine).

Figure 4

Temporal evolutions of the antidepressant-like effects caused by HCE75, the standardized flavonoid mixture containing equivalent doses of rutin, hesperidin, quercetin, and quercitrin and the fixed-ratio combination of rutin and hesperidin (75:21.5, w/w) on mice in TST. The mice were tested 0, 0.5, 1, 2, 3, 4, and 8 h after administering (p.o.) with the vehicle (physiological saline with 10% Tween 80), HCE75 (400 mg/kg), standardized flavonoid mixture (8 mg/kg), and rutin/hesperidin fixed-ratio combination (8 mg/kg). Data are expressed as mean ± S.E.M. (n = 10). Data were analyzed using one-way ANOVA for multiple comparisons, followed by post hoc Student–Newman–Keuls test. *p < 0.05, **p < 0.01, and ***p < 0.001 compared with the control group (tested 1 h after vehicle treatment).

Figure 5

Effects of HCE75, the standardized flavonoid mixture containing equivalent rutin, hesperidin, quercetin, and quercitrin doses and the fixed-ratio combination of rutin and hesperidin (75:21.5, w/w), on (A) immobility time in TST and (B to D) their respective line crossings in OFT. The mice were tested 1 h after administration (p.o.) of the vehicle (physiological saline with 10% Tween 80), fluoxetine (20 mg/kg), HCE75 (20, 200, 400, 800, and 1600 mg/kg), standardized flavonoid mixtures (0.4, 4, 8, 16, and 32 mg/kg), and the fixed-ratio rutin and hesperidin combination (0.4, 4, 8, 16, and 32 mg/kg). Data are expressed as mean ± S.E.M. (n = 10). Data were analyzed using one-way ANOVA for multiple comparisons, followed by post hoc Student–Newman–Keuls test. *p < 0.05, **p < 0.01, and ***p < 0.001 compared with the control group (vehicle); ##p < 0.01 and ###p < 0.001 compared with the positive reference group (fluoxetine).

Figure 6

Dose–response curves of the immobility time in TST with respect to the control groups. (A) The positive reference group (fluoxetine, 20 mg/kg, p.o.) response was 0 for the antidepressant-like effects of rutin, hesperidin, and the fixed-ratio combination of rutin/hesperidin (75:21.5, w/w). (B) The consequent isobolograms. Data are expressed as mean ± S.E.M. (n = 10).

Figure 7

Effects of HCE75 on monoamine neurotransmitter levels in the prefrontal cortex (A and C) and hippocampus (B and D) of mice without (A and B) or with TST (C and D). The mice of the groups unexposed to TST were decapitated and were subjected to brain surgery 66 min after administration (p.o.) of the vehicle (physiological saline with 2% Tween 80), fluoxetine (20 mg/kg), and HCE75 (20, 200, 400, 800, and 1600 mg/kg). The mice of the blank group were untreated, but similarly underwent brain surgery. The mice of the groups exposed to TST were decapitated and were subjected to brain surgery immediately after the animals were tested 60 min after treatment. Data are expressed as mean ± S.E.M. (n = 10). Data were analyzed using one-way ANOVA for multiple comparisons, followed by post hoc Student–Newman–Keuls test. ^x p < 0.05, ^y p < 0.01, and ^z p < 0.001 compared with the blank group; ^a p < 0.05, ^b p < 0.01, and ^c p < 0.001 compared with the vehicle groups exposed to TST.

Figure 8

Representative histopathological sections of the (A) liver, (B) kidney, and (C) spleen of the mice treated with HCE75 (5000 mg/kg, p.o.) once a day for 1 d (acute treatment) or 21 consecutive days (chronic treatment). The control group was not treated. Insignificant lesions were observed in the experiments. Hematoxylin–eosin stains, 400 × original magnification.