An Update on Oligosaccharides and Their Esters from Traditional Chinese Medicines: Chemical Structures and Biological Activities

A great number of naturally occurring oligosaccharides and oligosaccharide esters have been isolated from traditional Chinese medicinal plants, which are used widely in Asia and show prominent curative effects in the prevention and treatment of kinds of diseases. Numerous in vitro and in vivo experiments have revealed that oligosaccharides and their esters exhibited various activities, including antioxidant, antidepressant, cytotoxic, antineoplastic, anti-inflammatory, neuroprotective, cerebral protective, antidiabetic, plant growth-regulatory, and immunopotentiating activities. This review summarizes the investigations on the distribution, chemical structures, and bioactivities of natural oligosaccharides and their esters from traditional Chinese medicines between 2003 and 2013.


Introduction
Oligosaccharides and their esters, a significant group of phytochemical compounds, are widely distributed in the roots, rhizomes, stems, barks, leaves, aerial, and whole parts of medicinal plants. They not only serve as the energy storage components, but also play a vital role in the treatment of diseases. Before 2003, there have been a number of reviews and reports in respect to the isolation and structure elucidation of oligosaccharides and their esters from Chinese medicinal plants [1][2][3], but few biological activities such as cancer chemopreventive, and protein kinase C inhibitory activities had been reported [4][5][6]. With the development of isolation and identification techniques [7][8][9][10][11], a larger number of oligosaccharides and their esters have been endlessly identified from traditional Chinese medicines in the past decades. These compounds have a wide variety of structure types because of the assembly of different monosaccharide units, the combination of various linking styles and the existence of kinds of substituents. And more promising biological activities associated with some of the oligosaccharides and their esters have been discovered. In vitro and in vivo investigations have demonstrated that they displayed antioxidant, antidepressant, anti-inflammatory, neuroprotective, cerebral protective, antidiabetic, cytotoxic, antineoplastic, plant growthregulatory, and immunopotentiating activities, and so forth. This review aims to provide a systemic summary of the studies on the distribution, chemical structures and biological activities of naturally occurring oligosaccharides and their esters from traditional Chinese medicines in the past decades. Among these compounds, the number of oligosaccharide esters is much greater than that of oligosaccharides, and the disaccharide esters are a very valuable source of active compounds. This information may help readers understand the structure characteristics and therapeutic indications of oligosaccharides and their esters from traditional Chinese medicines and offer clues to the development of new drugs. Orobanchaceae, Acanthaceae, Rosaceae, Musaceae, Sparganiaceae, Leguminosae, Equisetaceae, Boraginaceae, Iridaceae, Alismataceae, Lamiaceae, Araliaceae, Rubiaceae, Oleaceae, Apocynaceae, Caryophyllaceae, Aspleniaceae and Trilliaceae, are rich in oligosaccharides and their esters. Oligosaccharides show diversified structures because of the type and the number of monosaccharides, as well as the position of glycosidic bonds. And oligosaccharide esters also display distinctive structural diversity largely owing to the number, type, and position of O-substituent units, including phenylpropanoid groups (e.g., coumaroyl, feruloyl, caffeoyl, sinapoyl, 3,4,5-trimethoxycinnamoyl, and cinnamoyl), benzoyl, p-methoxybenzoyl, and p-hydroxybenzoyl groups ( Figure 2). Moreover, the double bonds of phenylpropanoid groups possess trans and cis isomeric forms, of which the trans forms widely exist in nature. Hence, according to the number of monosaccharides and the characteristics of chemical structures, these oligosaccharide esters could be categorized into 7 large groups. (Table 1 and Figure 1) merely consist of various monosaccharides without O-substituents. In addition to the well-known sucrose, -D-glucopyranosyl(1 → 2)--D-glucopyranoside (1) was isolated from Camptosorus sibiricus [12]. The oligosaccharides of raffinose (3), stachyose (19), and verbascose (21), all of which belong to the Raffinose family, possess one, two or three galactopyranosyl units linked to sucrose and have been found in the rhizomes and roots of Alisma orientalis [13], Lycopus lucidus [14], Rehmannia glutinosa [15,16], Salvia miltiorrhiza [17], and Scrophularia ningpoensis [18]. Manninotriose (4) and verbascotetraose (5) consisting of galactopyranosyl units and a glucopyranosyl unit have been isolated from Alisma orientalis [13].

Periploca sepium
Root barks [35] 34  (Table 3) with a sucrose moiety possess six different types of fatty acid residues which attach to the 6 or 6 position of sucrose. These fatty acids include linoleic acid, palmitic acid, linolenic acid, myristic acid, hexadeca-7,10,13-trienoic acid, and hexadeca-7,10dienoic acid. The above sucrose fatty acid esters have been found in the rhizomes of Astragalus membranaceus and the roots of Equisetum hiemale.

Lignan-Derived Disaccharide Esters.
Six lignan-derived disaccharide esters ( Figure 4) contain a sucrose core esterified with different lignan residues covalently linked to the 3 and 6 positions of -D-fructofuranosyl unit and have been isolated from the whole parts of Trigonotis peduncularis [85] and the aerial parts of Eritrichium rupestre [86].

Phenylpropanoid-Derived Pentasaccharide Esters.
As shown in Table 6, oligosaccharide esters (160-179) possessing a skeleton of five sugar residues have been isolated from the roots of Polygala tenuifolia. The sugar residues are composed of two types of monosaccharides including fructofuranose and glucopyranose, which are esterified with acetyl, benzoyl, rhamnose-substituted/nonsubstituted coumaroyl, and rhamnose-substituted/nonsubstituted feruloyl groups. Other than that, a structure-complex oligosaccharide polyester shown in Figure 5, polygalajaponicose I (180), consisting of a pentasaccharide backbone esterified with feruloyl, coumaroyl, rhamnosyl-coumaroyl, acetyl, and benzoyl groups has been obtained from the roots of P. japonica [90].

Smilax riparia
Roots and rhizomes [59] Evidence-Based Complementary and Alternative Medicine 9 Scheme 2 antidepressant, cytotoxic, antineoplastic, anti-inflammatory, antidiabetic, plant growth-regulatory, neuroprotective, and cerebral protective activities. Lignan-derived disaccharide esters, phenylpropanoid-derived tetrasaccharide esters, and pentasaccharide esters with biological activities have not been reported. Aside from the isolated constituents, oligosaccharide mixtures from Rehmannia glutinosa, Panax ginseng, and Scrophularia ningpoensis were also reported to display diverse pharmacological activities, such as antidiabetic, immunopotentiating, enhanced memory, and antineoplastic activities. These active compounds and mixtures could serve as the valuable candidates to be developed as possible drugs for the treatment and prevention of diseases.

Antioxidant Activity.
The adverse effects of oxidative stress proposed to play significant roles in the pathogenesis of cardiovascular diseases, atherosclerosis, hypertension, cancer, diabetes mellitus, neurodegenerative diseases, rheumatoid arthritis, ischemia/reperfusion injury, and ageing have become an inevitable and serious issue [95,96]. Scientists have thus made great efforts to explore antioxidants from medicinal plants by using different kinds of assay methods, which include DPPH radical scavenging assay, hydroxyl radical scavenging assay, superoxide anion scavenging assay, and ABTS radical scavenging method [96]. Lapathosides C and D, hydropiperoside, vanicoside B, hidropiperosides A and B, lapathoside A, and diboside A were isolated from the Polygonum, Persicaria, and Fagopyrum genera belonging to the Polygonaceae family. The DPPH test revealed that free radical-scavenging activity of the isolated compounds termed lapathoside C (115), hydropiperoside (42), vanicoside B (111), and lapathoside D (38) increased in turn, and lapathoside D exhibited strongest scavenging ability with an IC 50 of 0.088 M [43]. Hidropiperosides A and B (117, 118) were reported to show obvious antioxidant response to DPPH radicals with the SC 50 values of 23.4 and 26.7 g/mL, respectively, while vanicoside E moderately exhibited the same activity with a SC 50 value of 49.0 g/mL [72]. Lapathoside A (114) and diboside A (116) just showed lower antioxidant activities with the SC 50 values of 199.48 and 165.52 M, respectively [73].
From the above studies, it can be concluded that oligosaccharide esters with antioxidant activities have been identified in the Polygonaceae, Liliaceae, Smilacaceae, and Arecaceae families. The results of the antioxidant assays show that the increased number of phenolic hydroxyl groups and acetyl groups could produce higher antioxidant activity. Fan et al. indicated that the increased number of phenylpropanoid groups was not beneficial to free radical scavenging activity [43]. Zhang et al. pointed out that oligosaccharide esters with feruloyl groups exhibited better antioxidant activities than those with coumaroyl groups [44].
Evidence-Based Complementary and Alternative Medicine 13

Antidepressant
Activity. The oligosaccharides obtained from the Morinda genus not only show specific antidepressant and antistress activities but also have no suppression or excitatory effects on central nervous system as well. What is more, they can be taken orally with little toxicity [21]. The inulin-type hexasaccharide (IHS) (9) from Morinda officinalis obviously exhibited cytoprotective activity, which contributed to the antidepressant effect, not only by providing the PC12 with protection against Cort-induced lesion with IHS 0.625 and 1.25 M, but also by reducing the Cort-induced [Ca 2+ ] overloading with IHS 2.5 and 10 M. IHS 5 and 10 M upregulated the nerve growth factor mRNA expression in Cort-induced PC12 cells [22]. Polygalatenosides A (181) and B (182) were isolated from the Polygala genus. They significantly inhibited the isotope-labeled RTI-55 binding to norepinephrine transporter protein with the IC 50 values of 30.0 and 6.04 M, respectively [91].
DISS and tenuifoliside A were isolated from the Polygala and Cynanchum genera. Liu et al. investigated the antidepressant effect of YZ ethanol extract based on the tail suspension test (TST) and forced swimming test (FST), which are the ease-of-use and widely-accepted models for estimating antidepressant activities in mice. The results indicated that YZ-50 fraction at a dose of 200 mg/kg was able to significantly decrease the immobility time in TST. Furthermore, YZ-50 possessed ability to inhibit corticosterone-induced injury of human neuroblastoma SH-SY5Y cells. What is more, DISS (123) and tenuifoliside A (44), two major compounds of YZ-50 fraction, showed effective protective response to the lesion in SY5Y cells [53].  the doses of 5, 10, and 20 mg/kg was also tested in chronically mild stressed rats. DISS was able to exhibit antidepressant activity by upregulating the expression of noradrenergicregulated plasticity genes including cell adhesion molecule L1, brain-derived neurotrophic factor, laminin, and cAMP response element binding protein factor in hippocampus [98]. DISS improved the reward reaction by increasing sucrose intake and obviously decreased the levels of serum cortisol, adrenocorticotrophic hormone, and corticotropinreleasing factor. Further, DISS played an enhanced role in the expression of mineralocorticoid receptor, together with glucocorticoid receptor mRNA [99].
Disaccharide esters and oligosaccharides mixture from the Liliaceae, Polygonaceae, Sparganiaceae, and Scrpophulariaceae families showed effective cytotoxic and antineoplastic activities. The study results indicated that feruloyl and acetyl groups play an important role in mediating cytotoxicity, which seems to be related to the substitution position of feruloyl groups. The feruloyl groups at C-6 or C-1 are vital for cytotoxicity. In addition, the increased number of acetyl groups could induce higher tumoricidal activity. 16 Evidence-Based Complementary and Alternative Medicine 3.4. Anti-Inflammatory Activity. Inflammation, an important basic pathological process, is a defense response of biopsy with vascular system to damage stimuli such as pathogens, impaired cells and tissues, and physical and chemical factors. However, if the process of inflammatory response cannot end normally when cell debris and pathogens were cleared, the biological defence response will become causative factor and bring about many diseases, such as diabetes, cardiovascular diseases, metabolic syndrome, and cancer [100,101].
Tenuifoliside A (44) from the Polygala genus exhibited strong anti-inflammatory effect not only by suppressing the production of NO, but also by reducing the production of iNOS, prostaglandin E2, cyclooxygenase-2, and proinflammatory cytokines through the inhibition of the mitogenactivated protein kinases pathway and NF-B pathway [102]. The anti-inflammatory activities of quiquesetinerviusides D (108) and E (109) from the Calamus genus were evaluated in RAW 264.7 cells. Both of them showed significant inhibitory effects against the production of LPS-stimulated NO with the IC 50 values of 9.0-29.5 M [70].
Six disaccharide fatty acid esters (132-137) were isolated from the Astragalus and Equisetum genera. The antiinflammatory effects of these isolated compounds have also been documented. The activation of NF-B could upregulate the expression of proinflammatory cytokines inducible nitric oxide synthase (iNOS) and tumor necrosis factor alpha (TNF-). The NF-B inhibitory activities of compounds 132-137 were tested in HepG2 cells stimulated with TNF-. All of these compounds could significantly restrain TNF--induced NF-B transcriptional activities with the IC 50 values of 4.4-24.7 M. Li et al. pointed out that olefinic bonds and the length of the fatty acid moieties contributed to the NF-B inhibitory activity. Furthermore, the inhibition increased significantly with the increase of the number of olefinic bonds on the aliphatic moiety [77]. These results may provide a scientific basis for the development of new anti-inflammatory agents.

Neuroprotective and Cerebral Protective Activities.
As we all know, glutamate works as a major excitatory amino acid neurotransmitter in the mammalian central nervous system and plays a crucial role in several physiological processes [103]. However, the accumulation of glutamate induces diverse acute and chronic neurodegenerative diseases, such as epilepsy, ischemic stroke, and Parkinson's disease, as well as Alzheimer's disease [104]. DISS (123) isolated from the Polygala genus exhibited neuroprotective effect against glutamate-induced SH-SY5Y neuronal cell damage. The in vitro test demonstrated that DISS (0.6, 6 and 60 mol/L) played a critical role in increasing cell viability, controlling lactate dehydrogenase and attenuated apoptosis ranging from 1.95% to 2.58% [105]. Tenuifoliside B (102) from the Polygala genus was able to significantly shorten the coma time of KCN-induced anoxia mice at the doses of 3 and 10 mg/kg, and it played an important role in ameliorating the scopolamine-induced impairment of performance in passive avoidance task in rats and enhancing the tremors induced by oxotremorine in mice. These results together demonstrated that tenuifoliside B possessed cognitive improving and cerebral protective effects [56].
3.6. Antidiabetic Activity. Diabetes mellitus, a chronic debilitating metabolic disease, is characterized by high blood glucose content and comprises three types termed type I, type II, and gestational diabetes [106]. Stachyose (19) extract (a part) from Rehmannia glutinosa obviously exhibited the activity of downregulating fasting plasma glucose level and partially keeping from hyperglycemia induced by adrenaline and glucose without obvious dose-dependent effect. Other than that, in vivo tests in rats induced by alloxan revealed that stachyose extract at the dose of 200 mg/kg significantly decreased blood-sugar level [15].
Diboside A, lapathosides C and D, vanicosides A and B, and hydropiperoside were isolated from the Fagopyrum, Polygonum, and Persicaria genera belonging to the Polygonaceae family. Diboside A (116) could potentially inhibitamylase activity with an IC 50 of 26.9 M and thus retard the starch digestion rate, which is helpful for diabetic individuals in controlling blood sugar level [107]. Lapathoside D (38) exerted stronger activity of -glucosidase inhibition with an IC 50 value of 0.113 mM than acarbose which was chosen as a positive drug for the treatment of type II diabetes [43]. Vanicoside B (111) was reported to have higher -glucosidase inhibitory activity with an IC 50 of 50.5 M than vanicoside A (110) with an IC 50 of 59.9 M because of the acetyl moiety of the latter possibly decreasing inhibitory activity of vanicoside A [71].
Fujimoto et al. investigated the inhibitory effects of mumeoses F-O (83-92) from the Prunus genus on aldose reductase and discovered that caffeoyl groups are crucial for the inhibitory effect on aldose reductase. And thus, mumeoses F, G, H, J, K, L, M and N (IC 50 = 22-77 M), with a coumaroyl group and acetyl groups, inhibited moderately aldose reductase from reducing glucose to sorbitol, which is associated with the chronic complications of diabetes [63,64].

Elicitors and Regulators.
Oligosaccharides are quite propitious for encoding biological information because of diverse monosaccharide units and complex molecular structures and they are therefore first described as biological   signals in plants [108]. Oligosaccharides from the cell wall fragments of plants and fungi are powerful signal molecules, such as the elicitors of plant defence response and the regulators of plant growth, and they are capable of exerting biological activities at exceedingly low concentrations [109]. Heptasaccharide (HS) (15) and octasaccharide (OS) (16) isolated from the Paris genus possessed plant growth-regulatory activities [29,30]. The two oligosaccharides significantly promoted the proliferation of Paris polyphylla var. yunnanensis roots at the doses of 2.5-20 mg/L. The octasaccharide had the most obvious effect on the growth of Panax japonicus var. major hairy roots at a dose of 30 mg/L, while the other had the most positive effect on saponin accumulation of Panax japonicus var. major hairy roots at a dose of 10 mg/L [29]. Similarly, Zhou et al. evaluated the stimulating effects of HS and OS on the root growth and saponin production of Panax ginseng hairy roots, which were induced from the plant roots infected with Agrobacterium rhizogenes strain A4. The results showed that there was a maximum effect on the hairy roots growth and saponin accumulation on day 10. Compared with control group, the root biomass dry weight was increased by more than 1.7-fold while the total saponin content of roots increased by more than 1-fold when these two oligosaccharides were added to the hairy root at a dose of 30 mg/L [30]. The above data illustrate that HS and OS could serve as the plant growth-regulators not only in their original species but also in others.
3.8. Immunopotentiating Activity. Macrophages are important targets of investigations on cytophagy, cellular immunity, and molecular immunology. Therefore, they are deemed to play a vital role in host defense comprising phagocytosis, proteolytic processing, pathogenic agent, apoptosis, cytokines production, and foreign antigens presentation [110]. The water-extracted oligosaccharides from Panax ginseng (WGOS) exhibited better immunopotentiating activity by increasing phagocytic function of macrophages and promoting NO, TNF-and reactive oxygen species production [110]. In addition, Wan et al. have obtained maltooligosaccharides (17, = 3∼8) and three oligosaccharides (2, 13, 14) from the Panax ginseng roots. The in vitro bioassay pointed out that WGOS could serve as efficacious stimulators of B and T lymphocytes [28]. These studies provided enlightenment that the mixture of oligosaccharides from Chinese herbal medicine exhibits significant effect on immune system.

3.9.
Others. Acetylcholinesterase (AChE) inhibitors show good therapeutic effects on myasthenia gravis, glaucoma, and Alzheimer's disease through reversible enzyme inhibition so as to increase the accumulation of acetylcholine in the synapse and then promote and prolong the function of acetylcholine. Vanicoside B (111) showed AChE inhibitory activity with an IC 50 of 0.062 mM, while hydropiperoside (42), and lapathosides C (115) and D (38) just exhibited weak enzyme inhibitory activity [43]. Wang et al. has explored low molecular mass carbohydrate polymer from Panax ginseng roots and obtained 30% ethanol elution (PGO) which included peptides and oligosaccharides (17, = 0∼5) identified as maltose, maltotriose, maltotetraose, maltopentaose, maltohexaose, and maltoheptaose. Pharmacological experiments revealed that PGO could significantly enhance the memory in scopolamine-induced memory deficit rats [27].
Cistanoside F (129) and kankanose (150) were isolated from the Cistanche, Paulownia, and Acanthus genera. Pharmacological experiments showed that cistanoside F and kankanose significantly exhibited vasorelaxant effects on the noradrenaline-induced contraction of thoracic aorta from rats [80].

Conclusion
Traditional Chinese medicine from natural kingdom plays an indelible role in the treatment of human diseases, and it has aroused the attention of those who have engaged in medicinal pharmaceutical chemistry. Therefore, scientists have made great contributions day after day to investigate the valid chemicals from traditional Chinese medicines. In the past decades, about 193 oligosaccharides and their esters have been identified from traditional Chinese medicinal plants. On the one hand, only a few oligosaccharides and their mixtures were investigated and just exhibited antidepressant, antineoplastic, antidiabetic, plant growth-regulatory, immunopotentiating, and enhanced memory activities. More exploratory work is still needed to excavate biological and pharmacological activities of oligosaccharides. On the other hand, oligosaccharide esters exhibited multi-advantageous activities. Bioassays have revealed that antioxidant, cytotoxic, antineoplastic, and anti-inflammatory activities are the most notable bioactivities. Of course, to search for the natural products with these activities is a hotpot in the contemporary drug research. Oligosaccharide esters provide a vast treasure trove for medical researchers. After considering the current studies, it should be taken as future directions to make more mechanism of action studies and clinical trials to further evaluate its potential as new drugs. Moreover, the structure-activity relationships discussed in this review will provide reference information for further exploring their relationships and continually discovering the new bioactive oligosaccharide esters.