METHODS: The fresh Azolla pinnata plant from Kuala Krai, Kelantan, Malaysia was used for crude extraction using Soxhlet and maceration methods. Then, the chemical composition of extracts and its structure were identified using GCMS-QP2010 Ultra (Shimadzu). Next, following the WHO procedures for larval bioassays, the extracts were used to evaluate the early 4th instar larvae of Aedes mosquito vectors.
RESULTS: The larvicidal activity of Azolla pinnata plant extracts evidently affected the early 4th instar larvae of Aedes aegypti mosquito vectors. The Soxhlet extraction method had the highest larvicidal effect against Ae. aegypti early 4th instar larvae, with LC50 and LC95 values of 1093 and 1343 mg/L, respectively. Meanwhile, the maceration extraction compounds were recorded with the LC50 and LC95 values of 1280 and 1520 mg/L, respectively. The larvae bioassay test for Ae. albopictus showed closely similar values in its Soxhlet extraction, with LC50 and LC95 values of 1035 and 1524 mg/L, compared with the maceration extraction LC50 and LC95 values of 1037 and 1579 mg/L, respectively. The non-target organism test on guppy fish, Poecilia reticulata, showed no mortalities and posed no toxic effects. The chemical composition of the Azolla pinnata plant extract has been found and characterized as having 18 active compounds for the Soxhlet method and 15 active compounds for the maceration method.
CONCLUSIONS: Our findings showed that the crude extract of A. pinnata bioactive molecules are effective and have the potential to be developed as biolarvicides for Aedes mosquito vector control. This study recommends future research on the use of active ingredients isolated from A. pinnata extracts and their evaluation against larvicidal activity of Aedes in small-scale field trials for environmentally safe botanical insecticide invention.
OBJECTIVE: This study investigates the chemical constituents, anti-proliferative, and apoptotic properties of C. nutans root extracts.
MATERIALS AND METHODS: The roots were subjected to solvent extraction using methanol and ethyl acetate. The anti-proliferative effects of root extracts were tested at the concentrations of 10 to 50 μg/mL on MCF-7 and HeLa by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay for 72 h. Morphological changes were observed under light microscope. Pro-apoptotic effects of root extracts were examined using flow cytometric analysis and RT-PCR. The chemical compositions of root extracts were detected using GC-MS.
RESULTS: The proliferation of MCF-7 cells was inhibited with the IC50 values of 35 and 30 μg/mL, respectively, for methanol and ethyl acetate root extracts. The average inhibition of HeLa cells was ∼25%. Induction of apoptosis in MCF-7 was supported by chromatin condensation, down-regulation of BCL2 and unaltered expression of BAX. However, only ethyl acetate extract caused the loss of mitochondrial membrane potential. GC-MS analysis revealed the roots extracts were rich with terpenoids and phytosterols.
DISCUSSION AND CONCLUSIONS: The results demonstrated that root extracts promote apoptosis by suppressing BCL2 via mitochondria-dependent or independent manner. The identified compounds might work solely or cooperatively in regulating apoptosis. However, further studies are required to address this.
METHODS: The enzyme was purified in two steps using affinity and size exclusion chromatography. Enzyme assays were performed using the malachite green assay and enzymatic product was identified using gas chromatography-mass spectrometry (GC-MS) analysis. Sequence analysis of PmSTS was performed using multiple sequence alignment (MSA) against plant sesquiterpene synthase sequences. The homology model of PmSTS was generated using I-TASSER server.
RESULTS: Our findings suggest that the recombinant PmSTS is mainly expressed as inclusion bodies and soluble aggregate in the E. coli protein expression system. However, the addition of 15% (v/v) glycerol to the protein purification buffer and the removal of N-terminal 24 amino acids of PmSTS helped to produce homogenous recombinant protein. Enzyme assay showed that recombinant PmSTS is active and specific to the C15 substrate FPP. The optimal temperature and pH for the recombinant PmSTS are 30 °C and pH 8.0, respectively. The GC-MS analysis further showed that PmSTS produces β-sesquiphellandrene as a major product and β-farnesene as a minor product. MSA analysis revealed that PmSTS adopts a modified conserved metal binding motif (NSE/DTE motif). Structural analysis suggests that PmSTS may binds to its substrate similarly to other plant sesquiterpene synthases.
DISCUSSION: The study has revealed that homogenous PmSTS protein can be obtained with the addition of glycerol in the protein buffer. The N-terminal truncation dramatically improved the homogeneity of PmSTS during protein purification, suggesting that the disordered N-terminal region may have caused the formation of soluble aggregate. We further show that the removal of the N-terminus disordered region of PmSTS does not affect the product specificity. The optimal temperature, optimal pH, Km and kcat values of PmSTS suggests that PmSTS shares similar enzyme characteristics with other plant sesquiterpene synthases. The discovery of an altered conserved metal binding motif in PmSTS through MSA analysis shows that the NSE/DTE motif commonly found in terpene synthases is able to accommodate certain level of plasticity to accept variant amino acids. Finally, the homology structure of PmSTS that allows good fitting of substrate analog into the catalytic active site suggests that PmSTS may adopt a sesquiterpene biosynthesis mechanism similar to other plant sesquiterpene synthases.
Methods: Chemical profiling of P. blanda was carried out using gas chromatography mass spectrometry (GCMS) followed by isolation of bioactive compounds by column chromatography. DPPH• and FRAP assays were used to evaluate antioxidant activity and the MTT assay was performed to estimate the cytotoxicity activity against three cancer cell lines, namely MCF-7, HL-60 and WEHI-3, and three normal cell lines, MCF10A, WRL-68 and HDFa.
Results: X-ray crystallographic data for peperomin A is reported for the first time here and N,N'-diphenethyloxamide was isolated for the first time from Peperomia blanda. Methanol and dichloromethane extracts showed high radical scavenging activity with an IC50 of 36.81 ± 0.09 µg/mL, followed by the dichloromethane extract at 61.78 ± 0.02 µg/mL, whereas the weak ferric reducing activity of P. blanda extracts ranging from 162.2 ± 0.80 to 381.5 ± 1.31 µg/mL were recorded. In addition, petroleum ether crude extract exhibited the highest cytotoxic activity against all the tested cancer cell lines with IC50 values of 9.54 ± 0.30, 4.30 ± 0.90 and 5.39 ± 0.34 µg/mL, respectively. Peperomin A and the isolated mixture of phytosterol (stigmasterol and β-sitosterol) exhibited cytotoxic activity against MCF-7 and WE-HI cell lines with an IC50 of (5.58 ± 0.47, 4.62 ± 0.03 µg/mL) and (8.94 ± 0.05, 9.84 ± 0.61 µg/mL), respectively, compared to a standard drug, taxol, that has IC50 values of 3.56 ± 0.34 and 1.90 ± 0.9 µg/mL, respectively.
Conclusion: The activities of P. blanda extracts and isolated compounds recorded in this study underlines the potential that makes this plant a valuable source for further study on anticancer and antioxidant activities.
METHOD: This was an unmatched case-control study in which children with ASD were recruited from an autism early intervention center and typically developed (TD) children were recruited from government-run nurseries and preschools. Urine samples were collected at home, assembled temporarily at study locations, and transported to the laboratory within 24 h. The Al concentration in the children's urine samples was determined using inductively coupled plasma mass spectrometry (ICP-MS).
RESULT: A total of 155 preschool children; 81 ASD children and 74 TD children, aged 3 to 6 years, were enlisted in the study. This study demonstrated that ASD children had significantly higher urinary Al levels than TD children (median (interquartile range (IQR): 2.89 (6.77) µg/dL versus 0.96 (2.95) µg/dL) (p 1, p