Quantification of human amniotic fluid constituents by high resolution proton nuclear magnetic resonance (NMR) spectroscopy

We have investigated the ability of high-resolution proton NMR spectroscopy to provide a biochemical constituent screening of human amniotic fluid (AF). Proton NMR spectra were obtained at 300 MHz on AF from patients undergoing amniocentesis in the mid-trimester. Only AF from normal pregnancies (normal fetal karyotype, normal a-fetoprotein levels, normal birth outcome) was used in this study. The AF supernatant was lyophilized and resuspended in deuterated water containing 0.1 mm phosphate buffer and 6.02 mm disodium maleate. Identification of low molecular weight compounds was confirmed by two-dimensional NMR spectra (primarily correlated spectroscopy, or COSY) and standard addition techniques. A broad profile of compounds were ‘NMR visible’ in a single proton spectrum, including creatinine, glucose, organic acids (acetate, citrate, and lactate) and several amino acids (alanine, histidine, leucine, phenylalanine, tyrosine and valine). The proton spectrum was unaffected by prior freezing/thawing of AF samples. We were able to quantify compounds by comparison with an added concentration standard (maleate) at concentrations as low as 30 μm. Good agreement with literature values based on other analytical techniques was obtained.     

Detection of low-grade mosaicism in fetal cells isolated from maternal blood

Recovering and analysing fetal erythrocytes from maternal blood is being pursued for non-invasive prenatal genetic diagnosis. We report the observation of 46, XY/47, XXY mosaicism in fetal cells from a woman whose first-trimester chorionic villus sampling (CVS) initially showed only 46, XY. Only after exhaustive (500 cells) analysis were four XXY cells found in cultured villi.     

Chorionic villus culture for prenatal diagnosis of chromosome defects: Reduction of the long-term cultivation time

We report in detail two series of chorionic villus cultivation for prenatal chromosomal diagnosis. Chorionic villi were sampled from both first- and second-trimester pregnancies. One hundred cultures were treated with trypsin–EDTA for 2 h and collagenase overnight, (method A) and 100 were treated with trypsin–EDTA for 1 h and collagenase for 2 h (method B). Using short-term enzymatic digestion, the cultivation time was reduced from 14 days to 6 days. Sufficient amounts of metaphases of good quality were present in 93 per cent of primary cultures harvested in situ, whereas enough metaphases of sufficiently good quality were in most cases present only after subcultivation of the cultures using method A. The decrease in cultivation time obtained is probably due to a higher yield of viable cells in monocellular suspension, an increased attachment efficiency, and a more rapid attachment of single cells (within 24 h).     

Microbial degradation of usnic acid in the reindeer rumen

Reindeer (Rangifer tarandus) eat and utilize lichens as an important source of energy and nutrients in winter. Lichens synthesize and accumulate a wide variety of phenolic secondary compounds, such as usnic acid, as a defense against herbivores and to protect against damage by UV-light in solar radiation. We have examined where and to what extent these phenolic compounds are degraded in the digestive tract of the reindeer, with particular focus on usnic acid. Three male reindeer were given ad libitum access to a control diet containing no usnic acid for three weeks and then fed lichens ad libitum (primarily Cladonia stellaris) containing 9.1 mg/g DM usnic acid for 4 weeks. Usnic acid intake in reindeer on the lichen diet was 91–117 mg/kg BM/day. In spite of this, no trace of usnic acid or conjugates of usnic acid was found either in fresh rumen fluid, urine, or feces. This suggests that usnic acid is rapidly degraded by rumen microbes, and that it consequently is not absorbed by the animal. This apparent ability to detoxify lichen phenolic compounds may gain increased importance with future enhanced UV-B radiation expected to cause increased protective usnic acid/phenol production in lichens.     

Green roof stormwater retention: effects of roof surface, slope, and media depth

Urban areas generate considerably more stormwater runoff than natural areas of the same size due to a greater percentage of impervious surfaces that impede water infiltration. Roof surfaces account for a large portion of this impervious cover. Establishing vegetation on rooftops, known as green roofs, is one method of recovering lost green space that can aid in mitigating stormwater runoff. Two studies were performed using several roof platforms to quantify the effects of various treatments on stormwater retention. The first study used three different roof surface treatments to quantify differences in stormwater retention of a standard commercial roof with gravel ballast, an extensive green roof system without vegetation, and a typical extensive green roof with vegetation. Overall, mean percent rainfall retention ranged from 48.7% (gravel) to 82.8% (vegetated). The second study tested the influence of roof slope (2 and 6.5%) and green roof media depth (2.5, 4.0, and 6.0 cm) on stormwater retention. For all combined rain events, platforms at 2% slope with a 4-cm media depth had the greatest mean retention, 87%, although the difference from the other treatments was minimal. The combination of reduced slope and deeper media clearly reduced the total quantity of runoff. For both studies, vegetated green roof systems not only reduced the amount of stormwater runoff, they also extended its duration over a period of time beyond the actual rain event.