We have explored the method of extraction and purification of cyclic-peptide extract (CPE) from Pseudostellaria heterophylla (Miq.) Pax. (Taizishen, TZS), characterized the structure about cyclic-peptide compounds and investigated the biological activity of CPE attenuating chronic obstructive pulmonary disease (COPD) in rats. The CPE from TZS was obtained by ethyl acetate, petroleum ether, hot water extraction, and alcohol-precipitation. Cyclicpeptide structures were distinguished using ultra-high performance liquid chromatographyquadrupole time-of-flight tandem mass spectrometry (UPLC-Q-TOF-MS/MS). Rats were induced by solid combustibles smoke (SCS) for the COPD model, and the anti-COPD activity of CPE was detected using lung airway resistance and dynamic lung compliance, as well as pulmonary tissue hematoxylin and eosin (HE) staining. The relevant inflammatory cytokines were assayed by enzyme-linked immunosorbent assay (ELISA). CPE obtained from TZS contained 12 cyclic-peptide constituents; the purity was up to 92.94%. CPE (200, 400, or 500 mg/kg/day) was given to SCS-induced COPD model rats orally for 15 days. The results showed that in rats given CPE (400 mg/kg/day) there was a sharp fall in lung airway resistance but a rise in dynamic lung compliance. The image analysis of lung tissue sections suggested that CPE could decrease the degree of alveolar destruction (p <0.05), alleviate lung inflammation, increase alveolar space, and improve the infiltration of inflammatory cells. CPE was found to reduce the levels of TNF-a, but increase IL-10, adjusting multiple cytokines in rat serum; the TLR4 mRNA, MyD88 mRNA and AP-1 mRNA levels, the expressing levels of p-JNK, p-p38 and p-TAK1 protein were significantly down regulated in rat alveolar macrophages. CPE intervention could improve the pulmonary ventilation function on COPD rats, which may be related to its effect in inhibiting the abnormal activation of the TLR4-MyD88-JNK/p38 pathway. This is the first report that the CPE of TZS lessens the severity of COPD episodes. The new preparation process of CPEs implements the anticipated goal, which is to refine CPE and actualize quality control.

Polysaccharide (PF40) has been recognized as a main bioactive substances inPseudostellaria heterophylla (Miq.). The current study explored the potential protective effects of PF40 on immune system in mice with cyclophosphamide-induced immunosuppression. The mice were intragastric administered PF40 at the dosage of 100, 200 or 400 mg/kg once daily for 30 days, On the 24th and 25th day, the additional intraperitoneal injection of PF40 (50 mg/kg) were administered. The results showed that PF40 enhanced the cell-mediated immunity via improvements in macrophage phagocytosis, splenocyte proliferation, NK cell activity and delayed type hypersensitivity. Equally, it improves humoral immunity through promoting the formation of serum hemolysin.Moreover, PF40 maintain the immune balance of splenic lymphocytes and altered the intestinal physiological status in Cyp-induced mice. PF40 regulates the intestinal microbiota by restoring the relative abundance of Odoribacter and Mucispirillum and reducing the relative abundance of Sporosarcina, Yaniella, and Jeotgalicoccus in Cypintervened mice. The findings suggested that PF40 might be a promising natural functional foods for reducing chemotherapy-induced immunosuppression.

This study aimed to investigate the Pseudostellaria heterophylla polysaccharides (PF40) physicochemical and antidiabetic characteristics. The ultraviolet–visible (UV) spectra, Fourier transform infrared radiation (FT-IR) spectra, nuclear magnetic resonance (NMR) spectra, zeta potential, surface characteristics, and conformational and thermal stability properties of PF40 were characterized. X-ray diffraction (XRD) and scanning electron microscopy (SEM), combined with Congo red test, revealed that PF40 powder has mainly existed in amorphous form with triple-helix conformation. The single-molecular structure of PF40 exhibited a multi-branched structure extending from the center to the periphery by scanning probe microscopy (SPM) scanning. The monosaccharide residue of PF40 was an α-pyranoid ring and exhibits good stability below 168 °C. Experimental studies on antidiabetic characteristics found that PF40 could significantly improve STZ-induced intestinal mucosal damage and reduce the apoptosis of villus epithelial cells. PF40 combined with metformin could significantly improve the symptoms of insulin resistance in type 2 diabetes mellitus (T2DM) rats, the molecular mechanism might be through inhibiting the expression of RORγ protein and increasing Foxp3 protein in the jejunum of T2DM rats, and then restoring the STZ-induced imbalance of T helper 17(Th17)/ regulatory T cells (Treg) cells, thereby maintaining intestinal immune homeostasis. Results identified in this study provided important information regarding the structure and antidiabetic characteristics of Pseudostellaria heterophylla polysaccharides, which can contribute to the development of Pseudostellaria heterophylla polysaccharides for industrial purposes in the future.

Polysaccharides are abundant in natural resources and perform numerous physiological functions. Polysaccharide structures often lack chromophore groups; thus, current analytical methods cannot distinguish polysaccharide metabolites in the body or polysaccharide prototypes in biological samples. Thus, the measurement of polysaccharides in blood, bodily fluid, and cell-culture medium is difficult. Our early-stage research resulted in the isolation of two homogeneous polysaccharides from Pseudostellaria heterophylla, PHP0.5MSC-F and PHPH-1-2, which have anti-hyperglycemia and insulin resistance improvement effects for type 2 diabetes. In this study, the reducing terminal sugars of PHP0.5MSC-F and PHPH-1-2 were labeled with 2-aminobenzamide (2-AB) to prepare novel fluorescent probes for HPLC-coupled fluorescence detection (HPLC-FLD). Quantitative analysis was performed in reference to T40, and the detection limit for PHP0.5MSC-F was found to be 8.84 μg/mL with a linear range of 29.45–683.28 μg/mL. In reference to T70, the detection limit for PHPH-1-2 was found to be 13.89 μg/mL with a linear range of 46.29–462.76 μg/mL. This method was used to measure the bidirectional transport of polysaccharides across caco-2 cells from apical to basolateral (AP→BL) or from basolateral to apical (BL→AP) directions and to evaluate the polysaccharide bioavailability. The drug absorption capacity was determined based on the apparent permeability coefficient (Papp), and the Papp values for the two polysaccharides were found to be greater than 1 × 10−6 cm/s, which suggests easy absorption. Regarding bidirectional transport, the AP→BL Papp values were greater than the BL→AP values; thus, PHP0.5MSC-F and PHPH-1-2 mainly underwent passive transference. The two membrane permeable polysaccharides were not P-gp efflux transporter substrates. The absorption mechanism of PHP0.5MSC-F complies with passive diffusion under a concentration gradient, whereas PHPH-1-2 mainly utilizes a clathrin-mediated endocytic pathway to enter caco-2 cells. This innovative HPLC-FLD method can help to track polysaccharide internalization in vitro and in vivo to facilitate cellular uptake and biodistribution exploration.