Sugars and sucrose transporters in pollinia of Phalaenopsis aphrodite (Orchidaceae).

The pollen grains of Phalaenopsis orchids are clumped tightly together, packed in pollen dispersal units called pollinia. In this study, the morphology, cytology, biochemistry, and sucrose transporters in pollinia of Phalaenopsis orchids were investigated. Histochemical detection was used to characterize the distribution of sugars and callose at the different development stages of pollinia. Ultra-performance liquid chromatography-high resolution-tandem mass spectrometry data indicated that P. aphrodite accumulated abundant saccharides such as sucrose, galactinol, myo-inositol, and glucose, and trace amounts of raffinose and trehalose in mature pollinia. We found that galactinol synthase (PAXXG304680) and trehalose-6-phosphate phosphatase (PAXXG016120) genes were preferentially expressed in mature pollinia. The P. aphrodite genome was identified as having 11 sucrose transporters (SUTs). Our qRT-PCR confirmed that two SUTs (PAXXG030250 and PAXXG195390) were preferentially expressed in the pollinia. Pollinia germinated in pollen germination media (PGM) supplemented with 10% sucrose showed increased callose production and enhanced pollinia germination, but there was no callose or germination in PGM without sucrose. We show that P. aphrodite accumulates high levels of sugars in mature pollinia, providing nutrients and enhanced SUT gene expression for pollinia germination and tube growth.

Using electrospray-assisted laser desorption/ionization mass spectrometry to characterize organic compounds separated on thin-layer chromatography plates.

Electrospray-assisted laser desorption/ionization (ELDI), an ionization method that combines laser desorption and electrospray ionization (ESI), can be used under ambient conditions to characterize organic compounds (including FD&C dyes, amines, extracts of a drug tablet) separated in the central track on a thin-layer chromatography (TLC) plate coated with either reversed-phase C18 particles or normal-phase silica gel. After drying, the TLC plate was placed on an acrylic sample holder set in front of the sampling skimmer of an ion trap mass analyzer. The chemicals at the center of the TLC plate were analyzed by pushing the sample holder into the path of a laser beam with a syringe pump. The molecules in the sample spot were desorbed by continuously irradiating the surface of the TLC plate with a pulsed nitrogen laser. Then, the desorbed sample molecules entered an ESI plume where they were ionized through the reactions with the charged species (including protons, hydronium ions and their cluster ions, solvent ions, and charged droplets) generated by electrospraying a methanol/water solution. MS/MS analyses were also performed to further characterize the analytes. The detection limit of TLC/ELDI/MS is approximately 10(-6) M. This was evaluated by using FD&C red dye as the standard. A linear relationship was found for the calibration curve with the concentration of FD&C red dye ranged from 10(-3) to 10(-6) M.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for the detection of hemoglobins as the protein biomarkers for fecal occult blood.

In this study, we discover that hemoglobins (Hb), highly water-soluble globular proteins that are the most predominant proteins detected by matrix-assisted laser desorption ionization/time-of-flight (MALDI-TOF) mass spectrometry in blood, can be used as protein biomarkers for fecal occult blood (FOB). Hemoglobins were extracted from the feces with pure water and separated from the solids in feces through centrifugation. Singly charged molecular ions of Hb-related alpha chains (theoretical MW: 15 126) and beta chains (theoretical MW: 15 867) were detected by MALDI-TOF operated in linear mode using 4-hydroxy-alpha-cyanocinnamic acid (alpha-CHC) as the matrix (with a volumetric ratio of 1:1). The detection limit of FOB using this method is estimated to be lower than 0.1 microg blood per mg of feces, which is approximately 10 to 100 times lower than that of the conventional chemical approaches. The foods and dietary supplements that commonly interfere with the conventional chemical assays of FOB - such as animal blood food products and tablets containing iron and vitamin C - do not interfere with the detection of Hb biomarkers during MALDI-TOF analysis.

Diagnosis of occult blood in human feces using matrix-assisted laser desorption ionization/time-of-flight mass spectrometry.

BACKGROUND: Guaiac-based chemical tests may be the most cost-effective for routine screening of fecal occult blood (FOB). Unfortunately, such approaches still suffer from the unreliability of the incidence of false-positive and false-negative results in subjects who are on an unrestricted diet. We developed a rapid, automatic, and reliable detection method for FOB using matrix-assisted laser desorption ionization/time-of-flight mass spectrometry (MALDI-TOF MS). METHODS: The water-soluble alpha- and beta-chain hemoglobins (Hb), which are the most predominant proteins found in blood, were used as protein biomarkers for the diagnosis of FOB. The saturated 4-hydroxy-alpha-cyanocinnamic acid (alpha-CHC) solution was found to be the best MALDI matrix for Hb analysis. RESULTS: The MALDI-TOF MS results were consistent with those provided through conventional chemical analysis, however, the sensitivity for the detection of FOB when using MALDI-TOF MS is approximately 10-100 times higher. In addition, the materials that commonly interfere with the conventional chemical assaying of FOB do not interfere with the detection of the Hb biomarkers during MALDI-TOF MS analysis. CONCLUSIONS: We demonstrate that MALDI-TOF MS analysis of Hb biomarkers is an alternative approach toward the rapid diagnosis of FOB.

Characterization of the chemical components on the surface of different solids with electrospray-assisted laser desorption ionization mass spectrometry.

In this study we demonstrate that electrospray-assisted laser desorption ionization (ELDI) mass spectrometry (MS) can be used to rapidly characterize major chemical components on the surfaces of different solids under ambient conditions. The major chemical components in (a) dried milks with different fat contents, (b) different color-regions of a painting, (c) the thin coating on a compact disc, (d) drug tablets, and (e) porcine brain tissue were rapidly characterized as protonated molecules [M+H](+) or sodiated molecules [M+Na](+) by ELDI-MS with minimum sample pretreatment. The ionized ions of synthetic polymer and dye standards were detected directly from dried sample solutions using either positive or negative ion mode. Further structural information for the FD&C Red dye was obtained through tandem mass spectrometric (MS/MS) analysis using an ion trap mass analyzer attached to the ELDI source.