Optimization extraction, structural features and antitumor activity of polysaccharides from Z. jujuba cv. Ruoqiangzao seeds

Highlights

• Extraction yield of polysaccharides from Chinese jujube seeds was optimized by RSM.

• Two fractions were obtained by DEAE-52 column fractionation.

• Molecular structure, morphology and antitumor mechanisms of ZJSPs-1 were analyzed.

• ZJSPs-1 was in aggregates of dense honeycombed and spherical like structures.

• ZJSPs-1 showed a strong inhibition effect on HeLa cells and induced cells apoptosis.

Abstract

As a discarded byproduct of jujube (Z. jujuba), its seeds contain various biological components. Response surface methodology was used to optimize ultrasound-assisted heating extraction conditions of polysaccharides from Z. jujuba cv. Ruoqiangzao seeds (ZJSPs). The optimal parameters were as follows: temperature 83.1 °C, time 100 min, ultrasonic power 140 W, and water-material ratio 33.5 mL/g, allowed a maximum yield of 1.97%. One main fraction (ZJSPs-1) was successfully purified by ion-exchange and gel-permeation chromatography. With a molecular weight of 342 kDa, ZJSPs-1 was composed of arabinose, glucose, xylose, galactose and rhamnose. The presence of pyranose-form sugars as well as both α- and β-configurations was validated by FT-IR and ¹H NMR. TG/DSC indicated that ZJSPs-1 has a favorable thermal stability. XRD suggested that ZJSPs-1 exhibited both crystalline and amorphous portions. ZJSPs-1 was in aggregates of homogenous-dense honeycomb like structures observed by SEM and AFM. ZJSPs-1 possessed antitumor activities against HeLa cells in a dose- and time-dependent manner at 24 and 48 h with IC₅₀ of 164.6 and 87.1 μg/mL, respectively. Fluorescent microscopic and flow cytometry revealed that ZJSPs-1 inhibited the proliferation of HeLa cells through apoptosis. Z. jujuba seeds are good sources for antitumor polysaccharides in pharmaceutical and food industries.

Introduction

Jujube (Ziziphus jujuba Mill.), known as “Chinese dates”, is both an important fruit and Traditional Chinese Medicine (TCM) consumed in southwest Europe and Southeast Asia regions [1]. Nowadays, jujube is widely cultivated in the middle and lower reaches of the Yellow River of China [2]. It has reported that 1.5 million ha of jujube's cultivated area obtain the annual production of 450,000 t [3]. As a TCM, jujube fruits have been considered to be beneficial to spleen, stomach and heart functions [4]. In both ancient and modern China, jujube fruits, which are a common ingredient in many TCMs, play an irreplaceable role in both “food therapy” and “medical diet therapy” of TCMs. Undeniably, it is extensively consumed as functional food and medical materials in clinical nutrition therapy because it has a high content of polysaccharides, vitamins, polyphenols and minerals [1,3,[5], [6], [7]]. Because of high nutritional and pharmacological values, jujube fruits have gained increasing popularity over the last several decades internationally, leading to more than one billion people consumption in Asia [[8], [9], [10]].

The most common use of whole jujube fruits (besides fresh fruits) is to produce “jujube soup” thought boiling in Chinese traditional diets. More importantly, whole jujube fruits are frequently being included in a Chinese herbal formulated decoction in pharmacological industry, which is aiming to enhance the medicinal values, to facilitate the absorption and/or to reduce the toxicity of individual herbs [11,12]. However, the nutritional value of its seeds in “jujube decoctions” has not attracted much attention. Moreover, due to the widespread consumption of jujube fruits, the byproducts derived from its leaves, roots and seeds are considered to be a potential and valuable source for biological compounds [[13], [14], [15], [16], [17]]. Specially, jujube fruits contain exceeding 45% (on a dry matter basis) seeds. Its seeds are considered as a good source of valuable compounds such as polysaccharides, polyphenols, essential oils and protein [[18], [19], [20], [21]]. It is worth noting that jujube seeds had antioxidant and anxiolytic effect [13,22]. More importantly, Han et al. [23] found that the antioxidant activity of jujube seeds extract was significantly higher than that of its pulp. Additionally, jujube seeds could also be used as functional absorbents [16,24].

Nowadays, great attention has been paid on polysaccharides from TCMs because of their high bioactivities and relatively low toxicity [25]. It has reported that polysaccharides structure and bioactivities are greatly diverse among different Z. jujuba varieties [1,6,[26], [27], [28]]. Therefore, it is quite necessary to find new polysaccharides from different Z. jujuba varieties and their byproducts for functional food and pharmaceutical industries.

Ziziphus jujuba Mill. cv. Ruoqiangzao is a main cultivar of Ziziphus jujuba (Z. jujuba) (Fig. S1). Up to now, there is limited study on Z. jujube seeds as a good source of bioactive polysaccharides. Among various extraction techniques of polysaccharides, ultrasound-assisted extraction (UAE) has been recommended as a highly efficient method [29,30]. More importantly, Comparing with the traditional extraction methods, UAE combined with heating extraction (UAHE) tends to improve polysaccharides extraction yields and biological activities [[31], [32], [33]]. Consequently, the aim of this study were to probe UAHE conditions (i.e. extraction temperature, time, ultrasonic power and water-material ratio (W/M ratio)) of Z. jujuba cv. Ruoqiangzao seeds polysaccharides (ZJSPs) with response surface methodology (RSM)-based central composite design (CCD). Moreover, we investigated the structural properties and in vitro antitumor activities of ZJSPs to enhance the biological value of Z. jujuba seeds in “food therapy” and “medical diet therapy”.