Acclimation to UV-B radiation and visible light in Lactuca sativa involves up-regulation of photosynthetic performance and orchestration of metabolome-wide responses.

UV-B radiation is often viewed as a source of stress for higher plants. In particular, photosynthetic function has been described as a common target for UV-B impairment; yet as our understanding of UV-B photomorphogenesis increases, there are opportunities to expand the emerging paradigm of regulatory UV response. Lactuca sativa is an important dietary crop species and is often subjected to rapid sunlight exposure at field transfer. Acclimation to UV-B and visible light conditions in L. sativa was dissected using gas exchange and chlorophyll fluorescence measurements, in addition to non-destructive assessments of UV epidermal shielding (SUV ). After UV-B treatment, seedlings were subjected to wide-range metabolomic analysis using liquid chromatography hybrid quadrupole time-of-flight high-resolution mass spectrometry (LC-QTOF-HRMS). During the acclimation period, net photosynthetic rate increased in UV-treated plants, epidermal UV shielding increased in both subsets of plants transferred to the acclimatory conditions (UV+/UV- plants) and Fv /Fm declined slightly in UV+/UV- plants. Metabolomic analysis revealed that a key group of secondary compounds was up-regulated by higher light conditions, yet several of these compounds were elevated further by UV-B radiation. In conclusion, acclimation to UV-B radiation involves co-protection from the effects of visible light, and responses to UV-B radiation at a photosynthetic level may not be consistently viewed as damaging to plant development.

Vitamin D levels in Jamaican patients with systemic lupus erythematosus.

Vitamin D has important effects on the immune system as it has been shown to exert antiproliferative and relative immunosuppressant effects. Low levels of this hormone may contribute to the immune activation in systemic lupus erythematosus (SLE) and other autoimmune diseases. Serum levels of 25-OH vitamin D were measured in 75 patients with SLE in Jamaica, using an enzyme-linked immunoassay. Correlations with clinical data and disease activity as determined by the BILAG index were determined. Of a total of 75 patients, 33 (44%) had vitamin D sufficiency with mean vitamin D level of 39.45 ng/ml (range, 30.35-58.16). Forty-two (56%) patients had either vitamin D deficiency or insufficiency, 30 (40%) had vitamin D insufficiency, mean 26.36 ng/ml (range, 20.26-29.88), and 12 (16%) had vitamin D deficiency, mean 16.07 ng/ml (range, 7.78-19.90). There was an overall negative relationship between the total disease activity score using the BILAG index and vitamin D levels, and this was influenced primarily by the relationship seen among the vitamin D-deficient subgroup. This was also impacted on by a patient population that was significantly skewed toward low disease activity. The negative association trended toward statistical significance. Vitamin D deficiency is prevalent among patients with SLE in Jamaica. A relationship between low serum levels of vitamin D and SLE activity may occur.

Expression, purification, and use as an antigen of recombinant sugarcane mosaic virus coat protein.

A high titre (1:10,000) antiserum was raised in a rabbit against the coat protein of sugarcane mosaic potyvirus (SCMV), by injecting a preparation of recombinant coat protein purified from a fusion protein expressed in E. coli. The fusion protein consisted of the MalE maltose binding protein (MBP) and the viral coat protein separated by the protease factor Xa cleavage site. The fusion protein was encoded by the plasmid pMAL-cCPM, which was constructed by cloning a modified coat protein gene to the 3' end of the MBP/factor Xa coding region. The coat protein gene was modified by site-directed mutagenesis so that the ATG start codon in the original construct was replaced by the codon AGC, deleting the NcoI restriction site (C/CATGG) and creating a unique Eco47III site (AGC/GCT). Endonuclease restriction with Eco47III resulted in a DNA fragment with GCT as the first three nucleotides. This triplet encodes alanine, which is the proposed N-terminal amino acid residue of the mature native coat protein. This modified coat protein coding region was ligated directly behind the nucleotide code for the amino acid recognition sequence for factor Xa. Expression was induced with IPTG and the recombinant fusion protein was extracted from the bacterial lysate by amylose resin column affinity chromatography and the two domains separated by factor Xa proteolysis. The coat protein was then purified from the maltose binding protein by ion exchange chromatography in buffer containing 6 M urea. A highly purified sample which contained 150 micrograms of both full-length and truncated coat proteins, was recovered from a litre of bacterial broth. The antiserum reacted with native coat protein in SCMV-infected sugarcane, and with recombinant coat proteins expressed in E. coli and sugarcane protoplasts with little or no cross-reaction with sugarcane proteins.

Monitoring fusion protein cleavage by capillary electrophoresis.

Capillary electrophoresis was successfully utilized to monitor the proteolytic cleavage of two fusion proteins; maltose binding protein-sugarcane mosaic potyvirus coat protein and maltose binding protein-paramyosin. The course of proteolysis by factor Xa was monitored with high sensitivity and virtually in real time. Capillary electrophoresis conditions were optimized to clearly resolve the fusion proteins, factor Xa, lysis products and buffer components. Rapid elucidation of time to complete digestion was possible and fate of released proteins could also be examined.

Analysis of serum methylmalonic acid for the determination of cobalt deficiency in cattle.

An improved method has been developed and validated for the determination of methylmalonic acid (MMA) in serum to determine cobalt deficiency in cattle. Serum samples were extracted with ethyl acetate and derivatised to form the propyl esters using 14% boron trifluoride-propanol derivatising reagent. Derivatised samples were analysed by capillary gas chromatography using split injection, a DB-17 30 m x 0.25 mm I.D. capillary column and flame ionisation detection. The detection limit for the method was 0.5 mumol/l and precision, determined by replicate analyses of spiked serum samples, was less than 2% relative standard deviation. When cobalt deficiency is defined as a MMA serum concentration of more than 2 mumol/l, the method was able to detect clinical deficiency of cobalt in animals with symptoms such as "coasty" coats and low weight gain.

Measurement of plasma oxalate by capillary gas chromatography and its validation by isotope dilution mass spectrometry.

A capillary gas chromatographic method for plasma oxalate and an isotope dilution mass spectrometric reference method, both using the same tert.-butyldimethylsilyl derivatives, are described. Similar reference ranges for both were found (4.93 +/- 1.48 and 4.70 +/- 1.44 mumol/l, respectively), together with a close correlation for results covering a wide range of oxalate concentrations.

Multiresidue method for determination of pesticides in kiwi fruit, apples, and berry fruits.

A rapid multiresidue procedure for fruits, based on methanol extraction, is presented. Water and highly water-soluble co-extractives are removed by partitioning the pesticides into toluene. Carbon-cellulose-Florisil cleanup is performed before gas chromatography on OV-225 with electron capture detection. A wide range of pesticides can be determined to 0.1 mg/kg without concentrating the extract. Gas chromatography of the crude toluene partition using SE-30 and an alkali flame ionization detector provides confirmatory data and allows detection of some carbamates. Dichlorvos, dimethoate, and acephate are determined directly in the methanol extract by flame photometric detection. High recoveries were obtained for 4 organochlorine insecticides, 13 organophosphorus insecticides, 2 synthetic pyrethroids, two N-methyl carbamates, and 10 fungicides. The method is economical of solvents, glassware, and time, and is recommended for routine surveillance of residue levels on fruit.