贮藏温度和气体状况对苹果果胶、多聚半乳糖醛酸酶变化的影响

在苹果贮藏前期温度和CO_2浓度由高到低同步变动形成双维变动气调(TDCA)。 前期高温期间,果实予以高浓度CO_2处理,可抑制高温促使早熟的不利效应,其保鲜效果优于冷库,接近传统气调贮藏,表现在果肉硬度下降平缓;原果胶转化为水溶性果胶的速度减慢;多聚半乳糖醛酸酶活性显著受抑。低浓度CO_2抑制高温效应的作用大为减弱。低温下,不同浓度CO_2作用差异不大,均优于对照。     

High performance liquid chromatography–tandem mass spectrometry for the determination of bile acid concentrations in human plasma

We report a sensitive and robust method to determine cholic acid (CA), chenodeoxycholic acid (CDCA), deoxycholic acid (DCA), lithocholic acid (LCA), ursodeoxycholic acid (UDCA), and their taurine- and glycine-conjugate concentrations in human plasma using liquid chromatography–tandem mass spectrometry. Activated charcoal was utilized to prepare bile acid-free plasma, which served as the biological matrix for the preparation of standard and quality control samples. Plasma sample preparation involved solid-phase extraction. A total of 16 bile acids and 5 internal standards were separated on a reverse column by gradient elution and detected by tandem mass spectrometry in negative ion mode. The calibration curve was linear for all the bile acids over a range of 0.005–5 μmol/L. The extraction recoveries for all the analytes fell in the range of 88–101%. Intra-day and inter-day coefficients of variation were all below 10%. A stability test showed that all the bile acids were stable in plasma for at least 6 h at room temperature, at least three freeze–thaw cycles, in the −70 °C or −20 °C freezer for 2 months, and also in the reconstitution solution at 8 °C for 48 h. Comparison of the matrix effect of bile acid-free plasma with that of real plasma indicated that the charcoal purification procedure did not affect the properties of charcoal-purified plasma as calibration matrix. This method has been used to determine the bile acid concentrations in more than 300 plasma samples from healthy individuals. In conclusion, this method is suitable for the simultaneous quantification of individual bile acids in human plasma.     

Disruption of Stard10 gene alters the PPARα-mediated bile acid homeostasis

STARD10, a member of the steroidogenic acute regulatory protein (StAR)-related lipid transfer (START) protein family, is highly expressed in the liver and has been shown to transfer phosphatidylcholine. Therefore it has been assumed that STARD10 may function in the secretion of phospholipids into the bile. To help elucidate the physiological role of STARD10, we produced Stard10 knockout mice (Stard10^−/−) and studied their phenotype. Neither liver content nor biliary secretion of phosphatidylcholine was altered in Stard10^−/− mice. Unexpectedly, the biliary secretion of bile acids from the liver and the level of taurine-conjugated bile acids in the bile were significantly higher in Stard10^−/− mice than wild type (WT) mice. In contrast, the levels of the secondary bile acids were lower in the liver of Stard10^−/− mice, suggesting that the enterohepatic cycling is impaired. STARD10 was also expressed in the gallbladder and small intestine where the expression level of apical sodium dependent bile acid transporter (ASBT) turned out to be markedly lower in Stard10^−/− mice than in WT mice when measured under fed condition. Consistent with the above results, the fecal excretion of bile acids was significantly increased in Stard10^−/− mice. Interestingly, PPARα-dependent genes responsible for the regulation of bile acid metabolism were down-regulated in the liver of Stard10^−/− mice. The loss of STARD10 impaired the PPARα activity and the expression of a PPARα-target gene such as Cyp8b1 in mouse hepatoma cells. These results indicate that STARD10 is involved in regulating bile acid metabolism through the modulation of PPARα-mediated mechanism.     

Thermodynamic and solution state NMR characterization of the binding of secondary and conjugated bile acids to STARD5

STARD5 is a member of the STARD4 sub-family of START domain containing proteins specialized in the non-vesicular transport of lipids and sterols. We recently reported that STARD5 binds primary bile acids. Herein, we report on the biophysical and structural characterization of the binding of secondary and conjugated bile acids by STARD5 at physiological concentrations. We found that the absence of the 7α-OH group and its epimerization increase the affinity of secondary bile acids for STARD5. According to NMR titration and molecular modeling, the affinity depends mainly on the number and positions of the steroid ring hydroxyl groups and to a lesser extent on the presence or type of bile acid side-chain conjugation. Primary and secondary bile acids have different binding modes and display different positioning within the STARD5 binding pocket. The relative STARD5 affinity for the different bile acids studied is: DCA > LCA > CDCA > GDCA > TDCA > CA > UDCA. TCA and GCA do not bind significantly to STARD5. The impact of the ligand chemical structure on the thermodynamics of binding is discussed. The discovery of these new ligands suggests that STARD5 is involved in the cellular response elicited by bile acids and offers many entry points to decipher its physiological role.     

贮藏温度和气体组成对苹果乙烯生物合成的影响

苹果果实贮藏在高变温条件下(10℃开始,以后降至0℃)的空气中,随着乙烯形成酶(EFE)和ACC合成酶活性的增高,果实内乙烯浓度迅速上升,ACC和MACC积累。低温(0℃)抑制乙烯生成,ACC和MACC也有积累,但保持较高的EFE,促进ACC合成酶不断上升。低温气调(CA)(3％CO_2,3％O_2)显著抑制乙烯生成和相应的酶活性。高变温情况下,先提高CO_2浓度至12％,然后,随温度变化CO_2降至6％,形成双变气调。双变气调对乙烯生物合成的抑制作用与CA相同。