Mass spectrometric evidence for the existence of oligomeric anthocyanins in grape skins

The fractionation of a grape skin extract by multilayer countercurrent chromatography coupled with step gradient elution allowed the preparation of a fraction almost devoid of free anthocyanins. This fraction appeared to be almost exclusively polymeric, as judged by liquid chromatographic-mass spectrometric (LC-MS) analysis, color-bleaching tests with sulfur dioxide, and thiolysis. Electrospray mass spectrometric analysis indicated that the pigmented material in this fraction was chiefly composed of direct condensation products of anthocyanin extending up to trimers. With regard to their linkages, the anthocyanin units in the oligomers were possibly linked by either an A-type (by both carbon-carbon and ether bonds) or B-type (by carbon-carbon bond) linkage, like proanthocyanidins. The terminal anthocyanin unit of the oligomers is consistently in the flavylium form but the extension units are in the flavan form for the A-type oligomers and in the flavene form for the B-type oligomers. Although their linkages still need to be defined rigorously, this is the first mass spectrometric evidence confirming the existence of anthocyanin oligomers in the grape skin extract.     

Behaviors of D- and L-lactic acids during the brewing process of sake (Japanese rice wine)

The amounts of D- and L-lactic acids during the brewing process of sake were determined by capillary electrophoresis using 2-hydroxypropyl-beta-cyclodextrin as a chiral selector. Because L-lactic acid, which prevents the growth of nonuseful microorganisms, is a raw material of sake, the ratio of L-lactic acid to total lactic acid is almost 1.0 at the initial stage of sake brewing. During brewing, the ratio decreased gradually and finally reached 0.39. Yeast (Saccharomyces cerevisiae) for sake brewing produced D-lactic acid, but not L-lactic acid in a culture medium. These results suggest that the decrease in the ratio of L-lactic acid to total lactic acid during sake brewing resulted in D-lactic acid production by yeast. The ratios in 18 brands of sake obtained commercially ranged from 0.23 to 0.78. The levels of D-lactic acid in sake (140-274 mg/L) were in a narrower range than those of L-lactic acid (61-461 mg/L). Although the D-lactic acid level in sake did not correspond to total lactic acid level, the L-lactic acid level correlated well with total lactic acid level (R(2) = 0.867). These results suggest that the ratio of L-lactic acid to total lactic acid in sake reflected the amount of L-lactic acid added at the initial stage of sake brewing.     

Effect of light intensity on diurnal differences in the supply of cell wall components to the innermost surface of developing S<Subscript>2</Subscript> layers of tracheids in <Emphasis Type="Italic">Cryptomeria japonica</Emphasis>

We investigated the effect of light intensity on diurnal differences in secondary wall formation of tracheids. Saplings of Cryptomeria japonica were grown in growth chambers with light intensity cycles set for 12-h high light: 12-h low light by combining two of four light intensity levels: 1.5, 2.8, 4.3, and 10.0 klx. Volumetric changes of differentiating cells were monitored by measuring the tangential strain on the inner bark surface, and the innermost surface of developing secondary walls of differentiating tracheids during the high-light and low-light periods was observed by field-emission scanning electron microscopy. Changes in the aspects of the innermost surface of developing secondary walls and the tangential strain corresponded to changes in the light intensity level. Cellulose microfibrils were clearly observed when the light intensity was high (10.0 or 4.3 klx) and the volume of differentiating cells was low, while abundant amorphous material was observed when the light intensity was lowest (1.5 klx) and the cells were turgid, regardless of the light intensity cycle. These results suggest that the diurnal periodicity in the supply of cell wall components to developing secondary walls is associated with changes in light intensity during the photoperiodic cycle.     

A model of anisotropic swelling and shrinking process of wood

To elucidate the origin of the shrinking anisotropy of wood during the drying process, as well as to begin to gain an understanding of the interaction between the moisture and the cell wall components, the shrinking process of a single wood fiber regarding water desorption was simulated by using an analytical model which was developed in the previous report (Part 1). Resulting data were compared with the experimental ones in this paper. The following conclusions were obtained: (1) The matrix substance, as a skeleton in the secondary wall, tends to shrink isotropically. However, the cellulose microfibrils, as a rigid framework of the cell wall, almost did not shrink at all due to the water desorption. As result, wood shrinks anisotropically during a drying process. The microfibril angle in the S2 layer is one of the most important factors related to the degree of shrinking anisotropy of the wood while drying. (2) According to the simulation, the expansive strain caused in the matrix skeleton by the water sorption increases by 15% (= 150,000 micro-strains) from the oven-dried condition to the green condition. Based on this value, the moisture content at the fiber saturation point is calculated to be about 35%, which is close to the experimentally obtained one. These results give quantitative evidences that the hygroexpansion of the wood cell wall is controlled by the mechanism of the reinforced matrix hypothesis. Received: 28 July 1998     

Deletion in cysteine-rich region of LDL receptor impedes transport to cell surface in WHHL rabbit

The Watanabe heritable hyperlipidemic (WHHL) rabbit, an animal with familial hypercholesterolemia, produces a mutant receptor for plasma low-density lipoprotein (LDL) that is not transported to the cell surface at a normal rate. Cloning and sequencing of complementary DNA's from normal and WHHL rabbits, shows that this defect arises from an in-frame deletion of 12 nucleotides that eliminates four amino acids from the cysteine-rich ligand binding domain of the LDL receptor. A similar mutation, detected by S1 nuclease mapping of LDL receptor messenger RNA, occurred in a patient with familial hypercholesterolemia whose receptor also fails to be transported to the cell surface. These findings suggest that animal cells may have fail-safe mechanisms that prevent the surface expression of improperly folded proteins with unpaired or improperly bonded cysteine residues.     

Interleukin-1 stimulates the secretion of hypothalamic corticotropin-releasing factor

There is now evidence that the immune system, during times of infectious challenge, can stimulate the secretion of glucocorticoids, the adrenal steroids that mediate important aspects of the response to stress. Specifically, secretion of interleukin-1 (IL-1), a monocyte lymphokine secreted after infection, appears at least in part responsible for this effect. Glucocorticoids are secreted in response to a neuroendocrine cascade involving, first, the brain, then the pituitary, and finally the adrenal gland. In this report, human IL-1 is shown to activate the adrenocortical axis at the level of the brain, stimulating the release of the controlling hormone corticotropin-releasing factor (CRF) from the hypothalamus. Infusion of IL-1 induced a significant secretion of CRF into the circulation exiting the hypothalamus, whereas immunoneutralization of CRF blocked the stimulatory effect of IL-1 on glucocorticoid secretion. IL-1 appeared to have no acute direct stimulatory effects on the pituitary or adrenal components of this system. Furthermore, IL-1 did not cause a nonspecific release of other hypothalamic hormones. Thus, the lymphokine acts in a specific manner to activate the adrenocortical axis at the level of the brain; this effect appears to be unrelated to the known pyrogenic effects of IL-1 within the hypothalamus.     

Crystal structure of rhodopsin: A G protein-coupled receptor

Heterotrimeric guanine nucleotide-binding protein (G protein)-coupled receptors (GPCRs) respond to a variety of different external stimuli and activate G proteins. GPCRs share many structural features, including a bundle of seven transmembrane alpha helices connected by six loops of varying lengths. We determined the structure of rhodopsin from diffraction data extending to 2.8 angstroms resolution. The highly organized structure in the extracellular region, including a conserved disulfide bridge, forms a basis for the arrangement of the seven-helix transmembrane motif. The ground-state chromophore, 11-cis-retinal, holds the transmembrane region of the protein in the inactive conformation. Interactions of the chromophore with a cluster of key residues determine the wavelength of the maximum absorption. Changes in these interactions among rhodopsins facilitate color discrimination. Identification of a set of residues that mediate interactions between the transmembrane helices and the cytoplasmic surface, where G-protein activation occurs, also suggests a possible structural change upon photoactivation.     

Liver cancer: neonatal estrogen enhances induction by a carcinogen

A single injection of 100 micrograms of estradiol benzoate into newborn rats was followed after weaning by dietary treatment with one of two dosages of the carcinogen N-hydroxy-N-2-fluorenylacetamide. Autopsies 26 weeks later showed a higher incidence of liver cancer in male and, particularly, female rats injected with hormone than in controls. The weights of livers were greater but gonads were smaller in size in the estradiol groups. Endocrine and possibly centralnervous-system factors may play roles in formation of liver tumors.     

Metabolism of rotenone in vitro by tissue homogenates from mammals and insects

Hydroxylation of rotenone in vitro in the enzyme system composed of microsomes and reduced nicotinamide-adenine dinucleotide phosphate, and in living mice and houseflies, yields products tentatively identified as rotenolone I; rotenolone II; 8'-hydroxyrotenone; 6',7'-dihydro-6',7'-dihydroxyrotenone; two rotenolones of each of the last-mentioned two compounds; and uncharacterized polar materials. The toxicity of certain of these rotenoids to mice is of the same order as that of rotenone.     

Role of adaptor TRIF in the MyD88-independent toll-like receptor signaling pathway

Stimulation of Toll-like receptors (TLRs) triggers activation of a common MyD88-dependent signaling pathway as well as a MyD88-independent pathway that is unique to TLR3 and TLR4 signaling pathways leading to interferon (IFN)-beta production. Here we disrupted the gene encoding a Toll/IL-1 receptor (TIR) domain-containing adaptor, TRIF. TRIF-deficient mice were defective in both TLR3- and TLR4-mediated expression of IFN-beta and activation of IRF-3. Furthermore, inflammatory cytokine production in response to the TLR4 ligand, but not to other TLR ligands, was severely impaired in TRIF-deficient macrophages. Mice deficient in both MyD88 and TRIF showed complete loss of nuclear factor kappa B activation in response to TLR4 stimulation. These findings demonstrate that TRIF is essential for TLR3- and TLR4-mediated signaling pathways facilitating mammalian antiviral host defense.