Search

The outbreak of coronavirus disease 2019 (COVID-19) has spread across the world and was characterized as a pandemic. To protect medical laboratory personnel from infection, most laboratories inactivate the virus causing COVID-19, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), in clinical samples before testing. However, the effect of inactivation on the detection results remains unknown. Here, we used a digital PCR assay to determine the absolute SARS-CoV-2 RNA copy number in 63 nasopharyngeal swab samples and assess the effect of inactivation methods on viral RNA copy number., The outbreak of coronavirus disease 2019 (COVID-19) has spread across the world and was characterized as a pandemic. To protect medical laboratory personnel from infection, most laboratories inactivate the virus causing COVID-19, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), in clinical samples before testing. However, the effect of inactivation on the detection results remains unknown. Here, we used a digital PCR assay to determine the absolute SARS-CoV-2 RNA copy number in 63 nasopharyngeal swab samples and assess the effect of inactivation methods on viral RNA copy number. Viral inactivation was performed by three different methods: (i) incubation with the TRIzol LS reagent for 10 min at room temperature, (ii) heating in a water bath at 56°C for 30 min, and (iii) high-temperature treatment, including autoclaving at 121°C for 20 min, boiling at 100°C for 20 min, and heating at 80°C for 20 min. Compared to the amount of RNA in the original sample, TRIzol treatment destroyed 47.54% of the nucleocapsid protein (N) gene and 39.85% of open reading frame (ORF) 1ab. For samples treated at 56°C for 30 min, the copy number of the N gene and ORF 1ab was reduced by 48.55% and 56.40%, respectively. The viral RNA copy number dropped by 50 to 66% after heating at 80°C for 20 min. Nearly no viral RNA was detected after autoclaving at 121°C or boiling at 100°C for 20 min. These results indicate that inactivation reduced the quantity of detectable viral RNA and may cause false-negative results, especially in weakly positive cases. Thus, use of the TRIzol reagent rather than heat inactivation is recommended for sample inactivation, as the TRIzol reagent had the least effect on the RNA copy number among the tested methods.

In this study, crude polysaccharides from Porphyra haitanensis (CPP) were obtained by water extraction and alcohol precipitation. The CPP were separated and purified by DEAE-cellulose column chromatography to give five components (PP1, PP2, PP3, PP4, and PP5) whose physicochemical properties were characterized. The sulfate-group content (20.1%) of PP3 was higher than those of the other four components. PP3 was treated with H2O2-VC, and then dialyzed to obtain degradation products PP3-1 (102 kDa), PP3-2 (72 kDa), PP3-3 (39 kDa), and PP3-4 (20 kDa). The antioxidant activities of the degradation products and PP3 were assessed, which revealed that PP3-4 was significantly more active than the other four components. PP3-4 was purified by Sephadex G-100 to give PP3-4a. The molar ratio of galactose to 3,6-anhydro-α- L -galactose in PP3-4a was 10:1. The structure of PP3-4a was characterized by FT-IR and 1D/2D NMR spectroscopies. The results reveal that the structure of PP3-4a contains two →3) β- D -galactose (1→4) 3,6-anhydro-α- L -galactose (1 →, and → 3) β- D -galactose (1→4) α- L -galactose-6-S (1 → repeating structural units, with the latter structural units dominating and partially substituted with methoxy groups at some C-6 positions of the β- D -galactose and C-2 positions of 3,6-anhydro-α- L -galactose. The anomeric glycosidic bonds include both α and β types.

Traditional separation and purification process of Lycium barbarum polysaccharides (LBP) includes water extraction, alcohol precipitation, deproteinization and ion-exchange column chromatography, which is complicated and time-consuming. In our study, retentate LBP-I and dialysate LBP-O were obtained from LBP by water extraction, alcohol precipitation and deproteinization. LBP-I was separated by fractional precipitation and three fractions (LBP-I-1, LBP-I-2 and LBP-I-3) were obtained. The three fractions were further purified by gel permeation chromatography to LBGP-I-1, LBGP-I-2 and LBGP-I-3 with yields of 0.05%, 0.03%, and 0.19%, respectively, which are higher than yields by traditional method. The physicochemical properties, biological activities of LBGP-I-1, LBGP-I-2 and LBGP-I-3 were investigated. The results indicated that LBGP-I-1 (3.19 × 104 Da) consists of arabinose (21.95%), glucose (51.22%) and galactose (17.07%); LBGP-I-2 (2.92 × 104 Da) mainly consists of arabinose (19.35%), glucose (32.26%) and galactose (35.48%); LBGP-I-3 (9.12 × 104 Da) mainly consists of arabinose (48.15%) and galactose (44.44%). LBGP-I-1 and LBGP-I-2 were different from the components purified by traditional method. LBGP-I-3 could most significantly enhance macrophages NO, phagocytic capacity, and acid phosphatase. LBP-O exhibits the strongest anti-oxidant activities in vitro. These results provided a reference for applications of Lycium barbarum polysaccharides which would benefit the development of industry and agriculture.

Previous studies have shown that crude polysaccharides from the Lycium barbarum fruit could inhibit cancer cell growth, but the major effective constituents are yet to be identified. In this study, we compared the effects of L. barbarum fruit polysaccharide fractions on the growth of hepatoma cells (SMMC-7721 and HepG2), cervical cancer cells (HeLa), gastric carcinoma cells (SGC-7901), and human breast cancer cells (MCF-7). LBGP-I-3 showed stronger inhibitory effects on MCF-7 cells (cell viability of 48.96%) than SMMC-7721 (cell viability of 78.91%) and HeLa cells (cell viability of 55.94%), and had no effect on HepG2 and SGC-7901 cells. In addition, LBGP-I-3 had no inhibitory effect on normal liver cells (L02, cell viability of 115.58%). Investigation of the underlying mechanism suggested that LBGP-I-3 inhibited the growth of cancer cells by cell cycle arrest and apoptosis. LBGP-I-3 arrested the cell cycle at the G0/G1 phase, altered mitochondrial function, activated oxidative stress, and regulated the MAPK signaling pathway to induce apoptosis. Thus, LBGP-I-3 may be a potential functional food ingredient for the prevention of cancer without toxicity to normal cells in vitro. These results could help further elucidate the structure-activity relationship of L. barbarum fruit polysaccharides and functional food development.

The fruits of Lycium barbarum are considered medicinal foods with high nutritional value and bioactivity. In this study, we aimed to evaluate the effect of a crude L. barbarum polysaccharide (LBP) and two derived fractions, LBP-1 and LBP-2, on the lifespan of Drosophila melanogaster (fruit fly). The average lifespan of fruit flies was extended by supplementing their diet with either of the three LBP preparations. In vivo analysis of antioxidant activities detected increased superoxide dismutase (SOD) and catalase (CAT) activities and decreased malondialdehyde (MDA) levels. Dietary LBP supplements significantly reduced the mortality rate of fruit flies induced by paraquat and hydrogen peroxide. Importantly, the strongest anti-aging activity was exhibited by the LBP-2 fraction, containing arabinogalactan with a molecular weight of 9 × 104 Da. Further studies showed that the anti-aging activity of LBP was, at least in part, mediated by an age-related signaling pathway (MAPK, TOR, S6K) and the expression of longevity genes (Hep, MTH, and Rpn11).

We obtained four soluble acid xylan fractions AGP-III-A, AGP-III-B, AGP-III-C and AGP-III-D from the insoluble Artemisia sphaerocephala Krasch gum (ASKG) polysaccharide by weak alkali treatment combined with H2O2-Vc oxidative degradation. Activity studies showed that the degradation components could reduce the cell viability of several cancer cell lines in a concentration-dependent manner, especially 4-O-Methylglucuronoxylan AGP-III-C with specific molecular weight and branching degree significantly reduced cancer cells viability and induced HepG2 apoptosis, also caused mitochondrial membrane dysfunction upregulated ROS levels, and induced G0/G1 arrest in HepG2 cells by cell cycle assay. Further, AGP-III-C mediates apoptosis in HepG2 cells by upregulating MAPK phosphorylation. The structure of AGP-III-C was characterized by uronic acid reduction, permethylation with GC-MS, and 2D-NMR analysis. The structure of AGP-III-C had a linear (1→4)-linked β-Xylf residue backbone with one branched 4-O-Me-α-GlcAp attached to the main chain by a (1→2)-glycosidic bond at every two β-(1→4)-Xylf units.