Review Article
Pharmacological Aspects of Jujubes
Department of Chemistry and Chemical Technologies, University of Calabria, 87036 Rende (CS), Italy
INTRODUCTION
Jujube (Fig. 1), the fruit of Ziziphus jujuba Mill. (Rhamnaceae family) is widely distributed in many regions of Asia, Australia and the Mediterranean basin1. Jujube is also known as Chinese date, red date or tsao and it has been cultivated for more than 4000 years in China2. Jujube has been traditionally used fresh or processed (dried) as food, food additive and flavoring agent for thousands of years, due to its high nutritional values3. In addition, different parts of Ziziphus jujuba have been used (alone or as ingredient in more complex formulations) in Traditional Chinese Medicine (TCM) for calming down the mind and facilitate sleep and for the treatment of gastrointestinal problems, weakness, hepatitis, urinary troubles, skin infections, fever, pharyngitis, bronchitis, anemia and cancer4-9. Several studies have indeed confirmed that jujube displays many important biological properties, including antioxidant, neuroprotective, anticancer, anti-inflammatory, immunomodulatory, antiobesity, cardio, hepato and gastrointestinal protective activities, thus supporting the putative health benefits deriving from its consumption. This study deals with the pharmacological activities of jujubes. Results from in vitro and in vivo studies are discussed also and are summarized in Table 1 and 2, respectively. The chemical structures of the compounds responsible for jujube bioactivity are reported in Table 3.
IN VITRO STUDIES
Antioxidant and neuroprotective properties: Jujube extracts played a protective role in rat pheochromocytoma PC12 cells, a commonly employed model in the evaluation of neuroprotective effects, against tert-butyl hydroperoxide (tBHP)-induced cytotoxicity, stimulating the transcriptional factors of the antioxidant response element10.
Water extracts containing high amounts of cyclic nucleotides (cGMP and cAMP) and polysaccharides showed the highest cell protecting effect against tBHP-induced cell death. Among water extracts from fruits at different ripening stages, green jujubes containing a significantly higher amount of flavonoids [namely (-)-catechin, (-)-epicatechin, procyanidyn B2 and quercetin 3-O-rutinoside] showed superior cell protective effects than ripe fruits10. The brain benefits of jujube were further studied by investigating neuronal differentiation of PC12 cells11. Neurite outgrowth of PC12 cells was found to be induced after administration of a chemically standardized jujube water extract containing nucleobases (at least 80 μg g1 of dried extract), cGMP and cAMP (at least 150 μg g1) and flavonoids (quercetin 3-O-rutinoside, quercetin 3-O-galactoside, quercetin 3-O-β-D-glucoside and kaempferol 3-O-rutinoside, in a total amount of at least 35 μg g1). In addition, the expressions of neurofilaments (NFs), neuronal cell-specific cytoskeleton proteins, showed a concentration-dependent increase in jujube-treated cells, with inductions by ∼150% for NF68 and NF160. Jujube-induced neurite outgrowth and neurofilament expression were inhibited by pre-treatment with H89, a cyclic AMP-dependent and selective protein kinase A inhibitor. Moreover, the phosphorylation of cAMP responsive element binding protein on PC12 cells was induced by jujube extract and blocked by H89. The cAMP contained in jujube water extract was found to be a crucial ingredient in neuronal differentiation of PC12. However, despite its relatively high amount, jujube cAMP did not fully account for the displayed jujube-induced beneficial effects. The researchers speculated that other compounds, such as flavonoids could be partly responsible for the effects of jujube. Chen et al.11 investigated the expression of neurotrophic factors and antioxidant enzymes in cultured astrocytes treated with the aforementioned chemically standardized jujube water extract. Administration of jujube extract for 24 h significantly up-regulate mRNA expression of nerve growth factor, brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor in a concentration-dependent manner. Moreover, treatment with jujube water extract induced mRNA expressions of several important antioxidant enzymes in a concentration-dependent manner12. Zare-Zardini et al.13 investigated the effect of a new peptide (Snakin-Z) from jujube in inhibiting acetylcholinesterase (AChE) and butyrylcholinesterase (BChe) enzyme activity.
The AChE and BChE were significantly inhibited by Snakin-Z with IC50 values of 580 and 720 μg mL1, respectively.
Anticancer properties: Huang et al.14 investigated the anticancer activity of jujube in human hepatoma cells (HepG2) and found that jujube extracts decreased the viability of the cells. Chloroform fraction of the initial crude extract was the most effective, inducing a concentration-dependent effect on apoptosis, G1 arrest at a low concentration (100 μg mL1) and G2/M arrest at a higher concentration (200 μg mL1). The induction of apoptosis has been reported to be the mechanism through which jujube extracts exert anticancer activities in different human tumor cell lines, including HEp-2, HeLa and Jurkat cell lines15. The inhibitory effects of jujube extracts on breast cancer have been also investigated16. Fractions ZE2 and ZE4, containing triterpenic acids, exerted the highest antiproliferative effects on estrogen receptor alpha (ERα) positive MCF-7 (IC50 values of 7.64 and 1.69 μg mL1, respectively) and ERα negative SKBR3 (IC50 values of 6.21 and 3.70 μg mL1, respectively) human breast cancer cells. Moreover, ZE2 and ZE4 induced apoptosis in both cell lines and did not affect cell viability of both normal human fibroblasts BJ1-hTERT and nonmalignant breast epithelial MCF-10A cells16. The key role of some triterpenic acids present in jujube in inducing cytotoxicity in several tumor cell lines was already known. Lupane-type triterpenic acids bearing a p-coumaroyl moiety at the C-3 position showed a higher cytotoxicity than non-esterified triterpenic acids, suggesting that the esterification of the alphitolic acids with a p-coumaroyl group could be a key structural feature in inducing cytotoxic activity17. More recently, the proliferation of HepG2 and MCF-7 cells were strongly inhibited following the administration of triterpenic acids isolated from jujube, with IC50 values lower than 2.5 μg mL‾1 18. Betulinic acid isolated from sour jujube fruits was complexed with β-cyclodextrin (β-CD) to improve its solubility and study its anti-proliferative and apoptotic effects on MCF-7 cells19. The complex was found to inhibit the proliferation of cells in a concentration-dependent way, arresting cell cycle in the G2/M phase and inducing apoptosis via the mitochondria transduction pathway. In particular, the complex was found to significantly inhibiting B-cell lymphoma 2 (Bcl-2) expression and promoting Bcl-2-like protein 4 (Bax) expression, causing caspase-3 and caspase-9 cascade activation19. Two polysaccharide fractions isolated from jujube were found to inhibit the proliferation of melanoma cells in a concentration-dependent manner and to induce arrest at the G2/M phase, leading to the formation of an apoptotic body and to an increase of the activity20 of caspase-3 and caspase-9. Water extracts from several jujube varieties were investigated for their anticancer properties and one of them (Bombay) was found to possess moderate cytotoxicity against the Jurkat cell line with IC50 values of 375 μg mL1. However, it did not significantly induce cell death via apoptosis21. Jujube extracts from eight different growth stages (S1-8) inhibited the growth of HeLa cervical cancer cells in a concentration-dependent manner, while only the extracts from the early stages inhibited Hel299 normal lung and A549 lung cancer cell growth. The researchers correlated these latter findings with the content of certain flavonoids (quercetin 3-rutinoside and quercetin 3-robinobioside, mainly) and antioxidant activity that were found to decrease as the fruit matured22.
Anti-inflammatory and immunomodulatory properties: Jujube is one of the ingredients of a Chinese formulation (Shi Zao) traditionally used to alleviate the severe inflammatory and irritant effects of Euphorbia plants. Yu et al.23 provided a rationale for these anti-inflammatory properties, reporting that jujube extracts containing triterpenic acids strongly decreased the levels of Nitric Oxide (NO) and tumor necrosis factor (TNF)-α in rat peritoneal macrophages stimulated with Euphorbia kansui or prostratin, a phorbol ester isolated from Euphorbia fischeriana. Several studies report that jujube has immune stimulatory activity, mainly due to its polysaccharide content. Pectin (Ju-B-2) extracted from jujube and consisting of a polygalacturonan backbone with rhamnogalacturonan interspersed in the main chain induced a concentration-dependent proliferation of spleen cells. By contrary, a simple polygalacturonan (Ju-B-3) was inactive. The presence of rhamnogalacturonan and the side chains were suggested to be key factors in stimulating cell proliferation24. Polysaccharide fractions extracted from Z. jujuba cv. Jinsixiaozao (ZSP) were found to stimulate the proliferation of splenocytes and peritoneal macrophages25. Interestingly, pectin with a degree of esterification of 49% was abundant in the most active fraction (ZSP3c). Jujube extracts are also known to have anti-complementary activities. Some triterpenic acids belonging to the oleanane-type family exhibited significant anticomplementary activities with IC50 values ranging from 62.4-88.9 μg mL1. Interestingly, other triterpenes were inactive, suggesting that the oleanane pattern is a crucial structural feature in inhibiting the hemolytic activity of human serum against erythrocytes26. Jujube water extracts containing nucleobases, cyclic nucleosides and flavonoids11 have been shown to exert dual immune-modulating properties by regulating the expressions of pro-inflammatory cytokines in macrophages. Under normal conditions, mRNA expression of interleukin (IL)-1β, IL-6 and TNF-α were up-regulated after 24 h treatment with jujube extract in cultured RAW 264.7 macrophages. By contrast, the over-expression of IL-1β and IL-6, in lipopolysaccharide (LPS)-stimulated macrophages was abolished upon administration of the jujube water extract, at both mRNA and protein levels5. Jujube effect in stimulating hematopoietic function has been investigated, by evaluating the expression of erythropoietin in cultured Hep3B human hepatocellular carcinoma cells. Administration of the aforementioned jujube water extract11 stimulated erythropoietin expression in a concentration-dependent manner and up to ~100% of increase. Moreover, jujube water extract induced the transcriptional activity of the regulator for erythropoietin expression in Hep3B cells transfected with a plasmid containing hypoxia response element27.
Antiobesity properties: Jujube extracts have been shown to circumvent adipogenesis in 3T3-L1 preadiocytes. Lipid accumulation and glycerol-3-phosphate dehydrogenase activity were suppressed following administration of jujube extracts, while cell viability was unaffected. The major inhibitory effects were obtained in the case of the chloroform fraction, which was able to down-regulate the expression of key adipogenic transcription factors, including peroxisome proliferator-activated receptor (PPAR)-gamma and CCAAT enhancer binding proteins (C/EBPs)28.
Cardioprotective properties: Fujiwara et al.29 reported that triterpenes from jujube were able to inhibit foam cell formation induced by acetylated LDL in human macrophages. Among the compounds tested, triterpenic acids, including oleanonic, pomolic and pomonic acids were found as the most active; suggesting that the carboxylic group at C-28 is an important structural feature in inhibiting foam cell formation.
IN VIVO STUDIES
Antioxidant and neuroprotective properties: The neuroprotective properties of jujube extract (ZJE) were investigated in gerbils under conditions of ischemic damage. Neuronal nuclei-immunoreactive neurons were more abundant in the hippocampus of the ZJE-treated ischemia group than in the vehicle-treated ischemia group 4 days after ischemia/reperfusion (I/R). Superoxide dismutase (SOD) and brain-derived neurotrophic factor were higher in the ZJE-treated ischemia group than in the vehicle-treated ischemia group 4 days after I/R. In addition, the amount of hydroxynonenal, a marker of lipid peroxidation, was significantly lower in the ZJE-treated ischemia group than in the vehicle-treated ischemia group after I/R30. Pahuja et al.31 reported on the anti-epileptic properties of jujube extracts in rats. Administration of hydroalcoholic extract of jujube (HEZJ, containing spinosin and jujubosides as the major components) resulted in up to 100% protection against experimental seizure models of epilepsy in rats challenged with pentylenetetrazole (PTZ) or maximal electroshock (MES). In addition, the PTZ (or MES)-induced oxidative stress and decrease in cholinesterase activity were significantly reversed by HEZJ, leading to significant improvements in memory and learning. The memory dysfunction in Alzheimers disease has been associated with cortical cholinergic deficiency and loss of cholinergic neurons of the nucleus basalis of Meynert (NBM). A standardized jujube hydroalcoholic extract administration had repairing effects on memory and behavioral disorders produced by NBM lesions in rats, by activating choline acetyltransferase and may have beneficial effects in treatment of AD patients32. Total Phenolic Content (TPC) of the extract was 48.8 mg of gallic acid equivalent (GAE) per gram dried extract. Total flavonoid and flavonol contents were 9.1 and 8.0 mg of rutin equivalent (RE) per gram dried extract, respectively.
Anti-inflammatory and immunomodulatory properties: An hydroalcoholic extract of jujube (ZJ) was found to exert anti-inflammatory properties in both acute and chronic models of Inflammation in rats. The ZJ induced a dose-dependent reduction of edema induced by sub-plantar administration of carrageenan in rat hind paw. Moreover, ZJ significantly decreased the formation of granuloma tissue and nitrite/nitrate level induced by interscapular implantation of a sterile cotton pellet. The ZJ was found to contain jujubosides, flavonoids and terpenes but the bioactive compounds responsible for the reported effects were not investigated33. The polysaccharide fractions from Z. jujuba cv. Jinsixiaozao (ZSP) discussed earlier was also reported to enhance thymus and spleen indices in mice25. Also in this case, the most active fraction (ZSP3c) was rich in pectin with a degree of esterification of 49%.
Anti-allergic properties: Naik et al.34 evaluated the anti-allergic and anti-anaphylactic activity of the ethanolic extract of jujube in asthma and allergy animal models. Milk-induced eosinophilia and degranulation of mesenteric mast cells were significantly prevented by jujube extract. In addition, jujube extract inhibited acetylcholine as well as histamine-induced tracheal chain contraction and antigen-induced contraction of sensitized guinea pig ileum. A preliminary phytochemical analysis of the extract showed the presence of triterpenoids (betulinic acid, oleanolic acid, betulin and lupeol), alkaloids, carbohydrates, proteins and flavonoids.
Cardioprotective properties: Cheng et al.35 evaluated the protective effect of phenolics from Chinese jujube against Myocardial Injury (MI) in rats. Pre-administration of both free and bound phenolics from jujube peel (JPFP and JPBP, respectively) significantly inhibited the production of malondialdehyde (MDA) and the activities of Ca2+-ATPase and Mg2+-ATPase induced by isoproterenol (ISO) or aluminum chloride. In addition, the reduction of superoxide dismutase (SOD), glutathione peroxidase, creatine kinase, lactate dehydrogenase and Na+/K+-ATPase activities was circumvented upon treatment with JPFP or JPBP. Sun and Jiang investigated the effect of an undescribed jujube extract on oxidative injury in heart muscles of exhausted training rats. Their results showed that jujube extract was able to inhibit lipid peroxidation in heart and Bax expression, to increase the activity of antioxidant enzymes and Bcl-2 expression, thus improving heart functions36.
Gastrointestinal protective properties: A few studies have been carried out to provide a rationale for the anecdotal use of Chinese jujubes for the treatment of gastrointestinal symptoms. The effect of Water Soluble Carbohydrate Concentrate (WSCC) extracted from jujube and consisting of glucose, fructose, pectin polysaccharide and hemicellulose was investigated on several intestinal and fecal indices in a hamster model. The WSCC was found to effectively reduce gastrointestinal transit time and the amount of daily fecal ammonia in hamsters, thus allowing intestinal health maintenance37. Jujube polysaccharides, containing xylose (31.3%), glucose (23%), fructose (21.6%) and mannose (12.9%) as main monosaccharide units were found to significantly reduce intestine malondialdehyde (MDA) level and increase the activity of antioxidant enzymes in a rabbit model stimulated with I/R of the small intestine38.
Hepatoprotective properties: Jujubes are traditionally used for the treatment of hepatitis in Northern China. The hepatoprotective properties of jujube ethanolic extract (FZJ) on liver injury induced by carbon tetrachloride (CCl4) were investigated in male ICR mice8. Two biomarkers of hepatic injury in blood, namely alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were found significantly lower following the administration of FZJ at the dose of 200 mg kg1. Moreover, FZJ administration inhibited lipid peroxidation and increased the activities of endogenous antioxidant enzymes. Wang et al.39 investigated the effects of jujube heteropolysaccharides (ZSP), containing L-arabinose as the main monosaccharide unit, on CCl4-induced liver oxidative stress in mice. The activities of ALT, AST and lactic dehydrogenase in serum and the hepatic malondialdehyde level were significantly reduced following the administration of ZSP at a dose of 400 mg kg1. Moreover, ZSP-treated mice showed a better profile of hepatosomatic index and antioxidant system with normal activity of glutathione peroxidase and SOD in the liver. Zhao et al.40 investigated the effects of jujube polysaccharides (ZSPs) on fructose-induced insulin resistance and dyslipidemia in mice. The serum levels of glucose, insulin and dyslipidemia increased by 20% fructose water were significantly reduced upon administration of ZSP at a dose of 400 mg kg1. Hepatoprotective effect of ZSP against liver steatosis induced by a high-fructose diet was also confirmed by histopathological tests. These findings indicate that jujube polysaccharides may ameliorate insulin resistance and dyslipidemia in fructose-treated mice.
CONCLUSION
This study shows that jujube exerts a number of important pharmacological activities and can be therefore considered a valuable source of nutraceuticals. However, some of the reported effects of jujube are still anecdotal and have not been fully correlated with the occurrence of one or more specific bioactive compounds. In addition, the biological properties of jujube have been observed only in studies using in vitro and/or animal models. Human intervention studies are therefore indispensable to obtain reliable scientific evidence of the health benefits of jujube, in order to validate it as a healthy fruit.