Amyloid fibril formation by casein and fatty acid composition in breast milk of mastitis patients.

Mastitis can cause changes in the nutrient composition of breast milk, which may be harmful to both newborns and lactating mothers. In this study we preliminarily evaluated amyloid fibrils formation by casein and fatty acids (FA), as well as their potential relation with each other in the breast milk of mastitis patients. Six healthy volunteers and six mastitis patients were recruited from the Maternal and Child Health Care Hospital in Changchun were enrolled. Amyloid fibril content was assessed by thioflavin T fluorescence analysis, transmission electron microscope, circular dichroism, and proton nuclear magnetic resonance. FA contents were measured by gas chromatography. Healthy breast milk contained no amyloid fibrils but inflammatory breast milk did. Several FAs (hendecanoic acid, myristolenic acid, pentadecenoic acid, eicosatrienoic acid) differed significantly between the two groups (p < .05). The concentrations of the eicosatrienoic acid and eleven carbonic acids in the inflammatory groups were lower than those in the healthy groups, but the myristolenic acid and pentadecenoic acid were the opposite trend. Early detection of amyloid fibrils should be performed in lactating mothers with mastitis. Changes in FAs may reflect the importance of abnormal metabolism in amyloid fibril formation. PRACTICAL APPLICATIONS: The work preliminarily clarified the relationship between inflammation, fibril content, and fatty acid (FA) composition in breast milk. Healthy milk contained no amyloid fibril formed by casein but the inflammatory milk did. FAs were also significantly different between the two groups. Thus, an early determination of amyloid fibrils in milk should be considered for lactating women with mastitis to avoid the further malignant development. Additionally, the changes in FAs may reflect the importance of abnormal metabolism and oxidative pathways in amyloid fibril formation in the breast. Therefore, this study provided foundations for further investigation on the association between inflammation, fibril content and FA composition in breast milk.

Identification of ginsenoside metabolites in plasma related to different bioactivities of Panax notoginseng and Panax ginseng.

Although the chemical components of Panax notoginseng (PN) and Panax ginseng (PG) are similar, their bioactivities are different. In this study, the differential bioactivities of PN and PG were used as the research object. First, the different metabolites in the plasma after oral administration of PN and PG were analyzed using a UPLC-Q/TOF-MS-based metabolomics approach. Afterward, the metabolite-target- pathway network of PN and PG was constructed, and thus the pathways related to different bioactivities were analyzed. As a result, 7 different metabolites were identified in PN group, and 10 different metabolites were identified in the PG group. In the PN group, the metabolite N1 was related to hemostasis, N1 and N3 were related to inhibiting the nerve center, antihypertensive, and abirritation. The metabolites N1, N3, N4, N5, and N6 were related to liver protection. The results showed that the metabolites G1, G2, G3, G5, and G6 in PG group were related to heart protection, and G1, G2, G6, and G9 were related to increased blood pressure. There were 13 signaling pathways related to different biological activities of PN (8 pathways) and PG (5 pathways). These pathways further clarified the mechanism of action that caused the different bioactivities between PN and PG. In summary, metabolomics combined with network pharmacology could be helpful to clarify the material basis of different bioactivities between PN and PG, promoting the research on PN and PG.

Identification of metabolites in plasma related to different biological activities of Panax ginseng and American ginseng.

RATIONALE: Panax ginseng (PG) and American ginseng (AMG) are both medicinal plants of the Panax genus in the Acanthopanax family. Although PG and AMG have similar components of ginsenosides, there are many differences of their bioactivities. In this study, the biochemical mechanisms of different bioactivities of PG and AMG were explored by researching the differential metabolites in plasma after administration of each of PG and AMG. METHODS: In order to explore the material basis of differential bioactivities, two groups of mice were administrated orally with PG and AMG, and the method of metabolomics was used to identify the differential metabolites in plasma. Then network pharmacology was used based on the differential metabolites. Afterward, the metabolite-target-pathway network of PG and AMG was constructed; thus the pathways related to different bioactivities were analyzed. RESULTS: Through principal component analysis and orthogonal projections to latent structures discriminant analysis, there were 10 differential metabolites identified in the PG group and 8 differential metabolites identified in the AMG group. Based on network pharmacology, the differential metabolites were classified and related to differential bioactivities of PG and AMG. In the PG group, there were 6 metabolites related to aphrodisiac effect and exciting the nervous system, and 5 metabolites associated with raised blood pressure. In the AMG group, 5 metabolites were classified as having the effect of inhibiting the nervous system, and 6 metabolites were related to antihypertensive effect. CONCLUSIONS: This study explored the material basis of the differential biological activities between PG and AMG, which is significant for the research of PG and AMG use and to promote human health.

The metabolites and biotransformation pathways in vivo after oral administration of ocotillol, RT5 , and PF11.

Ocotillol, pseudo-ginsenoside RT5 (RT5 ), and pseudo-ginsenoside F11 (PF11 ) are ocotillol-type saponins that have the same aglycone structure but with different numbers of glucose at the C-6 position. In this study, the metabolites of ocotillol, RT5 , and PF11 in rat plasma, stomach, intestine, urine, and feces after oral administration were investigated by ultra-performance liquid chromatography coupled with time-of-flight mass spectrometry. The results showed that RT5 was easily biotransformed into metabolites in vivo, whereas PF11 and RT5 were difficult to be biotransformed. Hydrogenation, dehydrogenation, dehydration, deglycosylation, deoxygenation, hydration, phosphorylation, deoxidation, glucuronidation, and reactions combining amino acid were speculated to be involved in the biotransformation of ocotillol, RT5 , and PF11 . Based on the structural analysis of metabolites, it was deduced that hydrogenation, dehydration, deoxidation, and reactions combining amino acid occurred on the aglycone structure, whereas deglycosylation, hydration, and phosphorylation occurred on the glycosyl chain. Further, metabolites in plasma, urine, feces, and tissues were different: First, glucuronidation products were found in urine, stomach, intestine, and feces, but not in plasma. Second, the ocotillol prototype was not identified in urine samples. Third, the RT5 prototype was found in stomach, intestine, feces, and urine, but not in plasma.

Different absorption and metabolism of ginsenosides after the administration of total ginsenosides and decoction of Panax ginseng.

RATIONALE: Panax ginseng C.A. Meyer (PG), which contains polysaccharides and ginsenosides as the major bioactive components, has been used to promote health and treat diseases for thousands of years in China. Total ginsenosides were extracted from a decoction of Panax ginseng (GD), which included both ginsenosides and polysaccharides, and dissolved in water to obtain a total ginsenosides aqueous solution (TGAS). To study their absorption and metabolism, the pharmacokinetics (PK) and metabolites of ginsenosides in vivo were investigated after the administration of GD and TGAS. METHODS: Rat and mice plasma samples were collected after the administration of GD and TGAS. Ultra-performance liquid chromatography coupled with time-of-flight mass spectrometry was used with the UNIFI platform to identify metabolites in the plasma sample. The pharmacokinetic parameters were calculated using a noncompartmental method in the Drug and Statistics software package. RESULTS: Thirty ginsenoside metabolites were identified in mice plasma, of which only seven were found in the rat plasma after the administration of GD. The PK of ginsenosides Rb1 , Rc, and Rd were also determined after the oral administration of GD and TGAS and showed significant differences in the pharmacokinetic parameters. CONCLUSIONS: There was no difference in the biotransformation pathways after the oral administration of GD and TGAS, indicating that there was no influence of polysaccharides on the biotransformation of ginsenosides in vivo. However, the pharmacokinetic parameters were different after the administration of GD and TGAS, possibly because of the polysaccharides in GD. This study should be of significance in exploring the basis of PG bioactivities and lays the foundation for the further development of new drugs using PG.

The Influence of Ca2+ and Zn2+ on the Amyloid Fibril Formation by ß-Casein.

BACKGROUND: The amyloid fibril formation in different tissues or organs is related to amyloidosis. The Ca2+, Zn2+ and heparan sulfate (HS) are important elements and compositions in human body, which play a key role in regulating various physiological activities. Recently, there are increasing evidence suggest that they are closely linked to the amyloid fibril formation. OBJECTIVE: The effect of Ca2+ and Zn2+ on the amyloid fibril formation by ß-casein was investigated in the absence and presence of HS, which was significantly to explore the relationship between the concentration changes of Ca2+ and Zn2+ and amyloid fibril formation. METHODS: In this work, the influence of Ca2+ and Zn2+ on the ß-casein fibril formation in the absence and presence of HS was investigated by various methods of Thioflavin T fluorescence assay, transmission electron microscopy and intrinsic fluorescence measure. RESULTS: The results demonstrated that Ca2+ and Zn2+ promoted the ß-casein fibril formation. The effect of Ca2+ was greater than that of Zn2+. Meanwhile, the both metal ions had stronger effects when ß-casein was incubated with HS together. In addition, it was also observed that the microenvironment of ß-casein was changed because the intrinsic fluorescence peaks were red-shifted on the influence of Ca2+ and Zn2+. CONCLUSION: Ca2+ and Zn2+ were capable of promoting the ß-casein fibril formation in the both absence and presence of HS. This work set up the foundation for further researching of the amyloidosis pathogenesis and provided new insight for us to understand relationship between the inflammation and amyloidosis.

The effect of heparan sulfate on promoting amyloid fibril formation by ß-casein and their binding research with multi-spectroscopic approaches.

As a molecular chaperone, ß-casein is difficult to form amyloid fibrils under physiological conditions due to its chaperone activity. Heparan sulfate (HS) has drawn attention of technologists all over the word because of its relation to amyloid deposits in some amyloidosis diseases. For better understanding the relationship between the ß-casein and HS, the multi-spectroscopic studies were employed. The data of thioflavin T (ThT) binding assay, transmission electron microscopy (TEM) and circular dichroism (CD) demonstrated that HS promoted fibril formation by ß-casein in the amount and the growth speed. The results of steady-state UV-vis absorption spectra, fluorescence spectroscopy and fluorescence lifetime revealed that the ß-casein-HS complexes were formed and HS quenched the fluorescence of ß-casein by a static quenching mechanism. On the basis of fluorescence analysis, the value of binding constant was equal to 1.17 × 107 L mol-1 at 338.15 K and there was about one binding site between them. According to thermodynamic parameters obtained, it was deduced that a spontaneous reaction happened, and protein-ligand complex was stabilized by hydrogen bonds and hydrophobic interaction. Furthermore, using fluorescence resonance energy transfer (FRET) assay, the value of binding distance between HS and Trp143 of ß-casein was calculated to be 0.93 nm. Finally, on the basis of synchronous fluorescence experiment, the polarity increasing and hydrophobicity decreasing around Trp143 occurred during the period of fibril formation by ß-casein.