Extract of Xylopia aethiopica and its kaurene diterpene, xylopic acid, improve learning and memory in mice

Abstract

Background: Cognitive dysfunction, presenting as learning and memory impairment, is a common manifestation in many chronic diseases of the nervous system. Some of these diseases include depression, epilepsy, and Alzheimer's disease. To date, few drugs or medicinal products have shown ability to improve learning and memory deficits. Neuroprotection is one of the mechanisms by which memory could be improved. The extract of Xylopia aethiopica and its kaurene derivative, xylopic acid, have previously demonstrated neuroprotective effects in animal models. The aim of the present study was to investigate the effect of an extract of Xylopia aethiopica fruit and xylopic acid, on learning and memory using murine models.

Materials and methods: Unripe Xylopia aethiopica fruits were collected, dried, and extracted using 70% v/v ethanol. Xylopic acid was isolated from the fruits using petroleum ether, concentrated with ethyl acetate and then recrystallized with petroleum ether before purifying with ethanol (96%^v/_v). Institute of Cancer Research (ICR) mice received oral doses of the extract of Xylopia aethiopica (XAE; 30, 100 and 300 mg/kg), xylopic acid (XA; 30, 100 and mg/kg), citicoline (300 mg/kg), piracetam (300 mg/kg) or ketamine (30 mg/kg) and saline (vehicle). The animals were then taken through the Morris water maze test (MWM), spontaneous alternation Y-maze test (Y-maze), and novel object recognition test (NOR), to assess learning and memory.

Results: In the NOR test, XAE (30, 100 and 300 mg/kg) and XA (30, 100 and 300 mg/kg) increased the percentage exploration and recognition index (p = 0.0005 and p < 0.0001, respectively) when compared to both vehicle and ketamine groups. Similarly, doses of XAE and XA as used in the NOR test increased the percentage alternation in the Y-maze test. Although XAE and XA treatments decreased the latencies to find hidden platform in the MWM test, it was not significantly different from the vehicle group. However, this decrease in latency differed significantly when compared to the ketamine group. Interestingly, both XAE and XA treatments increased the percentage frequency to the target quadrant in the probe trial of the MWM. It is noteworthy that in all the three models used, both the extract and xylopic acid performed better than piracetam and citicoline, the reference drugs.

Conclusion: The ethanolic extract of Xylopia aethiopica fruit and xylopic acid improved exploratory learning and recognition memory, spatial working, recognition, and reference memories in the behavioral tests.

Keywords: AUC, Area under the curve; CTC, Citicoline; HPLC, High performance liquid chromatography; ICR, Institute of Cancer Research; KET, Ketamine; MWM, Morris water maze; Morris water maze; NOR, Novelty object recognition; Nootropic; Novelty object recognition; PCT, Piracetam; Spontaneous alternation Y-maze; TLC, Thin layer chromatography; VEH, Vehicle; XA, Xylopic acid; XAE, Extract of Xylopia aethiopica; Xylopia aethiopica; Xylopic acid; Y-maze, Spontaneous alternation Y-maze.

Fig. 1

Effects of XAE (30, 100 and 300 mg/kg) and XA (30, 100 and 300 mg/kg) treatment on the percentage time spent with the novel object in the NOR test. Data are presented as mean ± SEM (n = 8) for the time-course graphs (A, C) and analyzed by Two-way ANOVA followed by Bonferroni’s test. The total times spent with the new object are presented as the areas under the curve (AUCs) (B, D) (One-way ANOVA followed by Tukey’s multiple comparison test). Significantly different from the saline (VEH): *p < 0.05 **p < 0.01, ****p < 0.0001; significantly different from ketamine: ^††p < 0.01, ^†††p < 0.001 and ^††††p ^<0.0001. The recognition index is presented (E, F) (One-way ANOVA followed by Tukey’s multiple comparison test). Significantly different from the saline (VEH): *p < 0.05 **p < 0.01, ****p < 0.0001; significantly different from ketamine: ^†p < 0.05, ^††p < 0.01.

Fig. 2

Effects of XAE (30, 100 and 300 mg/kg) and XA (30, 100 and 300 mg/kg) treatment on the percentage attempted alternation in the blocked arm of the Y-maze in the spontaneous alternation Y-maze test measuring spatial working memory. Data are presented as mean ± SEM (n = 8) for the time-course graphs (A, C) and analyzed by Two-way ANOVA followed by Bonferroni’s test and their areas under the curve (AUCs) (B, D) (One-way ANOVA followed by Tukey’s multiple comparison test).Significantly different from the saline (VEH): *p < 0.05***p < 0.001, ****p < 0.0001; Significantly different from ketamine: ^††p < 0.01, ^†††p < 0.001 and ^††††p ^<0.0001.

Fig. 3

Effects of XAE (30, 100 and 300 mg/kg) and XA (30, 100 and 300 mg/kg) treatment on the percentage alternation in the previously blocked arm (i.e. opened) of the Y-maze in the spontaneous alternation Y-maze test measuring spatial recognition memory. Data are presented as mean ± SEM (n = 8) for the time-course graphs (A, C) and analysis of differences done by Two-way ANOVA followed by Bonferroni’s test. The areas under the curve (AUCs) (B, D) were analyzed by One-way ANOVA followed by Tukey’s multiple comparison test. Significantly different from the saline (VEH): *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001,; significantly different from ketamine: ^††p < 0.01, ^†††p < 0.001 and ^††††p < 0.0001.

Fig. 4

Effects of XAE (30, 100 and 300 mg/kg) and XA (30, 100 and 300 mg/kg) treatment on the percentage latency to locate hidden platform in the Morris water maze test. Data are presented as mean ± SEM (n = 8) for the time-course graphs (A, C) and analyzed by Two-way ANOVA followed by Bonferroni’s test and their areas under the curve (AUCs) (B, D) (One-way ANOVA followed by Tukey’s multiple comparison test). Significantly different from the saline (VEH): **p < 0.01; significantly different from ketamine: ^††p < 0.01 and ^††††p < 0.0001.

Fig. 5

Effects of XAE (30, 100 and 300 mg/kg) and XA (30, 100 and 300 mg/kg) percentage frequency of probe trial in the Morris water maze test measuring reference memory. Data are presented as mean ± SEM (n = 8) for (A) XAE and (B) XA groups. Significantly different from the saline (VEH): ***p < 0.001 and ****p < 0.0001. Significantly different from Ketamine: ^††p < 0.05, ^††p < 0.01 and ^††††p < 0.0001 (Analysis by differences by One-way ANOVA followed by Tukey’s multiple comparison test).