A New Class of 3′‐Sulfonyl BINAPHOS Ligands: Modulation of Activity and Selectivity in Asymmetric Palladium‐Catalysed Hydrophosphorylation of Styrene

Palladium‐catalysed monophosphorylation of (R)‐2,2′‐bisperfluoroalkanesulfonates of BINOL (R^F=CF₃ or C₄F₉) by a diaryl phosphinate [Ar₂P(O)H] followed by phosphine oxide reduction (Cl₃SiH) then lithium diisopropylamide‐mediated anionic thia‐Fries rearrangement furnishes enantiomerically‐pure (R)‐2′‐diarylphosphino‐2′‐hydroxy‐3′‐perfluoralkanesulfonyl‐1,1′‐binaphthalenes [(R)‐ 8ab and (R)‐ 8g–j ], which can be further diversified by Grignard reagent (RMgX)‐mediated CF₃‐displacement [→(R)‐ 8c–f ]. Coupling of (R)‐ 8a–j with (S)‐1,1′‐binaphthalene‐2,2′‐dioxychlorophosphine (S)‐ 9 generates 3′‐sulfonyl BINAPHOS ligands (R,S)‐ 10a–j in good yields (43–82%). These new ligands are of utlility in the asymmetric hydrophosphonylation of styrene ( 1 ) by 4,4,5,5‐tetramethyl‐1,3,2‐dioxaphospholane 2‐oxide ( 2 ), for which a combination of the chiral ligands with either [Pd(Cp)(allyl)] or [Pd(allyl)(MeCN)₂]⁺/NaCH(CO₂Me)₂ proves to be a convenient and active pre‐catalyst system. A combination of an electron‐rich phosphine moiety and an electron‐deficient 3′‐sulfone moiety provides the best enantioselectivity to date for this process, affording the branched 2‐phenethenephosphonate, (−)‐iso‐ 3 , in up to 74% ee with ligand (R,S)‐ 10i , where Ar=p‐anisyl and the 3′‐SO₂R group is triflone.     

Antifungal activity of Lactobacillus paracasei ssp. tolerans isolated from a sourdough bread culture

Lactic acid bacteria were isolated from four different sourdough bread cultures previously investigated for antifungal activity. A total of 116 isolates were obtained and screened for antifungal activity against a battery of molds. The most inhibitory isolate obtained was identified by API 50 CHL and 16s ribosomal RNA genotyping and found to be Lactobacillus paracasei ssp. tolerans. This isolate completely inhibited the growth of Fusarium proliferatum M 5689, M 5991 and Fusarium graminearum R 4053 compared to controls in a dual agar plate assay.     

Reduced toxicity of fumonisin B1 in corn grits by single-screw extrusion

Corn grits spiked with 30 microg/g fumonisin B1 and two batches of grits fermented with Fusarium verticillioides (batch 1 contained 33 microg/g, and batch 2 contained 48 microg/g fumonisin B1), which were extruded by a single-screw extruder with and without glucose (10%, dry weight basis) supplementation were fed to rats. Control groups were fed uncontaminated grits. Extrusion with glucose more effectively reduced fumonisin B1 concentrations of the grits (75 to 85%) than did extrusion alone (10 to 28%). With one exception, the fumonisin B1-spiked and fermented extrusion products caused moderately severe kidney lesions and reduced kidney weights, effects typically found in fumonisin-exposed rats. Lesions in rats fed the least contaminated grits (batch 1) after extrusion with 10% glucose were, however, significantly less severe and not accompanied by kidney weight changes. Therefore, extrusion with glucose supplementation is potentially useful for safely reducing the toxicity of fumonisins in corn-based products and studies to determine the optimal conditions for its use are warranted.     

Hypothalamic regulatory peptides and their receptors: Cytochemical studies of their role in regulation at the adenohypophyseal level

Hypothalamic regulatory peptides bind to specific receptors on target cells in the pituitary and control secretion. They in turn can be regulated at the pituitary level by steroid and peptide modulators. Affinity cytochemical techniques are important tools for the identification of specific target binding sites for these regulatory peptides. This presentation reviews the work in which potent, biotinylated ligands of gonadotropin releasing hormone (bio-GnRH), corticotropin releasing hormone (bio-CRH), and arginine vasopressin (bio-AVP) were applied to study the target cell responses. Bio-GnRH, bio-CRH, and bio-AVP bind to membrane receptors on specific anterior pituitary cells. Dual labeling for either gonadotropin or adrenocorticotropin (ACTH) antigens further identified the target cells. After 1–3 minutes, the label was in patches or capped on the surface. After 3 minutes, it was internalized in small vesicles and sent to receptosomes and vacuoles in the Golgi complex. Eventually the biotinylated peptides, or a metabolite, was found in the lysosomes (multivesicular bodies) and a subpopulation of secretory granules. The route and rate of uptake was similar to that described for the classical receptor-mediated endocytosis process. In contrast, intermediate lobe corticotropes internalized the bio-CRH in less than 1 minute. The route through the Golgi complex appeared to be bypassed. Instead the labeled peptide was in vesicles, on the membranes of scattered vacuoles, and in multivesicular bodies. Modulation of ligand binding by steroids showed that changes in receptor numbers correlated with changes in the number of cells that bound the ligand. In male rats, dihydrotestosterone reduced the percentage of GnRH-bound cells by 50%. Most of the reduction appeared in cells that stored luteinizing hormone (LH) antigens. In diestrous female rats, estradiol increased the percentage of bio-GnRH-bound cells. However, the steroid decreased the percentage of GnRH-bound cells in cells from proestrous rats. Glucocorticoids decreased the percentage of CRH-bound corticotropes in as little as 10 minutes. Potentiation of secretion by these ligands was correlated with increases in the percentage of ligand-bound cells. AVP pretreatment of corticotropes increased the percentage of cells that bound bio-CRH. It also increased the rate of receptor-mediated endocytosis of CRH and changed the route so that the Golgi complex was bypassed. This effect could be mimicked by activation of its second messengers (calcium and protein kinase C). Similarly, CRH pretreatment increased the percentage of corticotropes that bound AVP. Thyrotropin releasing hormone (TRH) pretreatment also increased the percentage of thyrotropes that bound AVP. Finally, calcium or sodium channel blockers altered CRH binding so that fewer cells were labeled. This binding by CRH was not dependent on extracellular calcium and tests with a calcium channel agonist showed that it was related to activation of calcium channels. To summarize, these affinity cytochemical studies have identified specific target cells in the pituitary for GnRH, CRH, and AVP. They have also identified heterogeneity in the population. They have demonstrated new information about the direct modulatory effects of steroids, ion channels, and neuropeptides on neuropeptide binding by subpopulations of these target cells.     

Physician's working diagnosis compared to the Euricterus Real Life Data Diagnostic Tool Trial in three jaundice databases: Euricterus Dutch, independent prospective and independent retrospective

BACKGROUND/AIMS: In the European Union Euricterus Project on (sub)Icterus proforma, the history and physical examination items were to be used for the physician's working diagnosis (PWD) and 'among others, for the development of the real life data electronic diagnostic tool, Trial. Trial delivers diagnosis probabilities based on Bayes' Theorem (B), completed by Trial Algorithm (TA). We wanted to compare the diagnostic accuracies (PWD and Trial probabilities as a percentage of the final diagnosis (FD) in a patient population) in 3 Dutch databases. METHODOLOGY: The inclusion criteria for both Euricterus and Trial were age > or = 16 and bilirubin > or = 20 mmol/l. Euricterus data gathering took place at the bedside on a proforma with (among other questions) 79 questions on history and physical examination as well as the diagnosis levels for the PWD (1 alternative possible) and FD (17 disease categories, dc). Trial was developed on the data of 7,104 Euricterus patients and its data-entry Demo has the same questions. It calculates the probability of each diagnosis of the 17 dc as a percentage, as each significant finding is encountered (BO, Bayesian Overall). It can simultaneously calculate the resemblance of the patient's signs and symptoms to each disease concomitantly (BV, Bayesian Vertical), and to any subset of a disease. Any probability is further tested for compatibility using TA, a subset of BV, delivering TA-PWD, TA-BO and TA-BV. The Trial test patients came from 3 databases: a Euricterus Dutch Patients Random Sample EDRS (n = 184, internal database) and 2 independent databases: prospective P (n = 80) and retrospective R (n = 152), totalling 416 patients. RESULTS: The accuracies of PWD and Trial showed no differences between the databases, and the results are therefore pooled (n = 416). With testing on the highest probability found, the PWD accuracy was 78%, TA-PWD 81%, TA-BO 74% and TA-BV 72%. The true FD's were mentioned (at any probability) in the PWD in 86%, TA-PWD in 92%, TA-BO in 94% and TA-BV in 91% of the patients. Testing only patients whose FD was "certain" or whose data were without omissions did not improve accuracy. Testing on probability > 95% improved BO and BV accuracy, but not TA-BO or TA-BV. CONCLUSIONS: The Physician's Working Diagnosis accuracy was approximately 80% and did not greatly improve after TA. The Trial TA-BO and TA-BV accuracies were only slightly less than the PWD. For well-trained physicians, Trial strengthens the physician's judgment, and for those less trained (or those to be trained), it delivers a (sub)icterus diagnostic disease probability at nearly consultant level.     

Modification of IgH PCR clonal analysis by the addition of sucrose and cresol red directly to PCR reaction mixes

We present the findings of a "new" sublethal MCA syndrome in three siblings, one female and two boys, the only children of healthy, non-consanguineous parents. In addition to prenatal growth retardation and early demise, they presented the same pattern of multiple malformations: relative microcephaly with bird-headed face, microphthalmos/coloboma iris/cloudy corneae, genital anomalies with hypospadias and cryptorchidism in the two males. Associated anomalies included: cardiac defects (2/3), unilateral cleft lip/cleft palate (1/3), anal stenosis (2/3) and unilateral renal agenesis (1/3).     

Rapid and parallel formation of Fe3+ multimers, including a trimer, during H-type subunit ferritin mineralization

Conversion of Fe ions in solution to the solid phase in ferritin concentrates iron required for cell function. The rate of the Fe phase transition in ferritin is tissue specific and reflects the differential expression of two classes of ferritin subunits (H and L). Early stages of mineralization were probed by rapid freeze-quench Mossbauer, at strong fields (up to 8 T), and EPR spectroscopy in an H-type subunit, recombinant frog ferritin; small numbers of Fe (36 moles/mol of protein) were used to increase Fe3+ in mineral precursor forms. At 25 ms, four Fe3+-oxy species (three Fe dimers and one Fe trimer) were identified. These Fe3+-oxy species were found to form at similar rates and decay subsequently to a distinctive superparamagentic species designated the "young core." The rate of oxidation of Fe2+ (1026 s(-1)) corresponded well to the formation constant for the Fe3+-tyrosinate complex (920 s(-1)) observed previously [Waldo, G. S., & Theil, E. C. (1993) Biochemistry 32, 13261] and, coupled with EPR data, indicates that several or possibly all of the Fe3+-oxy species involve tyrosine. The results, combined with previous Mossbauer studies of Y30F human H-type ferritin which showed decreases in several Fe3+ intermediates and stabilization of Fe2+ [Bauminger, E. R., et al. (1993) Biochem. J. 296, 709], emphasize the involvement of tyrosyl residues in the mineralization of H-type ferritins. The subsequent decay of these multiple Fe3+-oxy species to the superparamagnetic mineral suggests that Fe3+ species in different environments may be translocated as intact units from the protein shell into the ferritin cavity where the conversion to a solid mineral occurs.     

Transgenic mice expressing human ApoB95 and ApoB97. Evidence that sequences within the carboxyl-terminal portion of human apoB100 are important for the assembly of lipoprotein

The structural features of apolipoprotein (apo) B that are important for its covalent linkage to apo(a) to form lipoprotein(a) (Lp(a)) are incompletely understood. Although apoB100 cysteine 4326 is required for the disulfide linkage with apo(a), other structural features, aside from a single free cysteine residue, must be important for apoB's initial interaction with apo(a) and for facilitating the formation of the disulfide bond. To determine if sequences carboxyl-terminal to cysteine 4326 affect the efficiency of Lp(a) formation, we used "pop-in, pop-out" gene targeting in a human apoB yeast artificial chromosome to introduce nonsense mutations into exon 29 of the apoB gene. The mutant yeast artificial chromosomes, which coded for the truncated versions of human apoB, apoB95, and apoB97, were then used to express these mutant forms of apoB in transgenic mice. As judged by in vitro assays of Lp(a) formation, apoB95 (4330 amino acids) formed a small amount of Lp(a) but did so slowly. In contrast, apoB97 (4397 amino acids) formed Lp(a) rapidly, although not quite as rapidly as the full-length apoB100 (4536 amino acids). These results were supported by an analysis of double-transgenic mice expressing both human apo(a) and either apoB95 or apoB97. In mice expressing both apoB95 and apo(a), there was only a trace amount of Lp(a) in the plasma, and most of the apo(a) was free, whereas in mice expressing both apoB97 and apo(a), virtually all of the apo(a) was bound to apoB97 in the form of Lp(a). These results show that sequences carboxyl-terminal to apoB95 (amino acids 4331-4536) are not absolutely required for Lp(a) formation, but this segment of the apoB molecule, particularly residues 4331-4397, is necessary for the efficient assembly of Lp(a).     

Psychosocial, behavioral, and health factors related to menopause symptomatology

California Proposition 65 (Prop65) provides a mechanism by which the manufacturer may perform a quantitative risk assessment to be used in determining the need for cancer warning labels. This paper presents a risk assessment under this regulation for professional and do-it-yourself insulation installers. It determines the level of insulation glass fiber exposure (specifically Owens Corning's R-25 PinkPlus with Miraflex) that, assuming a working lifetime exposure, poses no significant cancer risk under Prop65's regulations. "No significant risk" is defined under Prop65 as a lifetime risk of no more than one additional cancer case per 100,000 exposed persons, and nonsignificant exposure is defined as a working lifetime exposure associated with "no significant risk." This determination can be carried out despite the fact that the relevant underlying studies (i.e., chronic inhalation bioassays) of comparable glass wool fibers do not show tumorigenic activity. Nonsignificant exposures are estimated from (1) the most recent RCC chronic inhalation bioassay of nondurable fiberglass in rats; (2) intraperitoneal fiberglass injection studies in rats; (3) a distributional, decision analysis approach applied to four chronic inhalation rat bioassays of conventional fiberglass; (4) an extrapolation from the RCC chronic rat inhalation bioassay of durable refractory ceramic fibers; and (5) an extrapolation from the IOM chronic rat inhalation bioassay of durable E glass microfibers. When the EPA linear nonthreshold model is used, central estimates of nonsignificant exposure range from 0.36 fibers/cc (for the RCC chronic inhalation bioassay of fiberglass) through 21 fibers/cc (for the i.p. fiberglass injection studies). Lower 95% confidence bounds on these estimates vary from 0.17 fibers/cc through 13 fibers/cc. Estimates derived from the distributional approach or from applying the EPA linear nonthreshold model to chronic bioassays of durable fibers such as refractory ceramic fiber or E glass microfibers are intermediate to the other approaches. Estimates based on the Weibull 1.5-hit nonthreshold and 2-hit threshold models exceed by at least a factor of 10 the corresponding EPA linear nonthreshold estimates. The lowest nonsignificant exposures derived in this assessment are at least a factor of two higher than field exposures measured for professionals installing the R-25 fiberglass insulation product and are orders of magnitude higher than the estimated lifetime exposures for do-it-yourselfers.     

Ureteropelvic junction injuries secondary to blunt abdominal trauma

PURPOSE: To describe the clinical and imaging findings of ureteropelvic junction (UPJ) injuries caused by blunt trauma. MATERIALS AND METHODS: In two children (aged 10 and 16 years) and eight adults (aged 23-82 years) with UPJ injuries, findings at computed tomography (CT) (n = 10), excretory urography (n = 6), and retrograde pyelography (n = 8) were retrospectively reviewed to identify the location and extent of contrast material extravasation. Clinical and follow-up data were correlated with radiologic findings. RESULTS: CT and urography played complementary roles in diagnosis. UPJ avulsion, defined as complete transection of the ureter with no filling of the ipsilateral ureter below the level of the UPJ, was diagnosed in four patients. UPJ laceration, defined as contrast material extravasation from the UPJ with contrast material in the ipsilateral ureter distal to the point of injury, was diagnosed in six patients. Medial perirenal contrast extravasation was seen in all 10 patients but failed to help differentiate UPJ avulsion from laceration. A distinctive pattern of contrast material extravasation at CT termed "circumrenal urinoma" was present in five patients and was found to be specific for UPJ injury. CONCLUSION: Medial perinephric contrast material extravasation was highly suggestive of UPJ injury. Demonstration of ureteral filling differentiated UPJ laceration from avulsion.