http://hdl.handle.net/10204/895 Tue, 21 May 2013 08:57:18 GMT 2013-05-21T08:57:18Z http://hdl.handle.net/10204/6675 Title: The short and long of it: shorter chromatographic analysis suffice for sample classification during UHPLC-MS-based metabolic fingerprinting Authors: Madala, NE; Tugizimana, F; Steenkamp, PA; Piater, LA; Dubery, IA Abstract: Ultra high-performance liquid chromatography hyphenated to mass spectrometry (UHPLC-MS) technologies has been widely applied in metabolomics, and the high resolution and peak capacity thereof are only some of the key aspects that are exploited in such and related fields. In the current study, we investigated if low resolution chromatography, with the aid of multivariate data analyses, could be sufficient for a metabolic fingerprinting study that aims at discriminating between samples of different biological status or origin. UHPLC-MS data from chemically-treated Arabidopsis thaliana plants were used and chromatograms with different gradient lengths were compared. MarkerLynxTM technology was employed for data mining, followed by principal component analysis (PCA) and orthogonal projections to latent structure discriminant analysis (OPLS-DA) as multivariate statistical interpretations. The results showed that, despite the congestion in low resolution chromatograms (of 5 and 10 min), samples could be classified based on their respective biological background in a similar manner as when using chromatograms with better resolution (of 20 and 40 min). This paper thus underlines that, in a metabolic fingerprinting study, low resolution chromatography together with multivariate data analyses suffice for biological classification of samples. The results also suggest that, depending on the initial objective of the undertaken study, optimisation in chromatographic resolution prior to full scale metabolomics studies is mandatory. Description: Copyright: 2012 Springer-Verlag. This is an ABSTRACT ONLY. The definitive version is published in Chromatographia, vol.76,(5-6), pp 279-285 Fri, 01 Mar 2013 00:00:00 GMT http://hdl.handle.net/10204/6675 2013-03-01T00:00:00Z http://hdl.handle.net/10204/6671 Title: Biotransformation of isonitrosoacetophenone (2-keto-2-phenyl-acetaldoxime) in tobacco cell suspensions Authors: Madala, NE; Steenkamp, PA; Piater, LA; Dubery, IA Abstract: Nicotiana tabacum cell suspensions, 2g wet wt/ml, rapidly took up 1 mM isonitrosoacetophenone (INAP), a plant-derived stress metabolite with anti-oxidative and anti-fungal properties, producing 40-hexopyranosyloxy-30-methoxyisonitrosoacetophenone in 54 %yield over 18 h. Unconverted INAP was at 33 lM. UPLC–MS/MS analyses with MassFragment software were used for metabolite identification. INAP had been hydroxylated at its meta- and para-positions as well as undergoing subsequent methoxylation and glycosylation. INAP is thus recognized by the enzymatic machinery of the phenylpropanoid pathway and is converted to a molecule with a substitution pattern similar to ferulic acid. Description: Copyright: 2012 Springer Netherlands. This is an ABSTRACT ONLY. The definitive version is published in Biotechnology Letters, vol.34(7), pp 1351-1356 Sun, 01 Jul 2012 00:00:00 GMT http://hdl.handle.net/10204/6671 2012-07-01T00:00:00Z http://hdl.handle.net/10204/6670 Title: Collision energy alteration during mass spectrometric acquisition is essential to ensure unbiased metabolomic analysis Authors: Madala, NE; Steenkamp, PA; Piater, LA; Dubery, IA Abstract: Metabolomics entails identification and quantification of all metabolites within a biological system with a given physiological status; as such, it should be unbiased. A variety of techniques are used to measure the metabolite content of living systems, and results differ with the mode of data acquisition and output generation. LC–MS is one of many techniques that has been used to study the metabolomes of different organisms but, although used extensively, it does not provide a complete metabolic picture. Recent developments in technology, for example the introduction of UPLC–ESI–MS, have, however, seen LC–MS become the preferred technique for metabolomics. Here, we show that when MS settings are varied in UPLC–ESI–MS, different metabolite profiles result from the same sample. During use of a Synapt UPLC–high definition MS instrument, the collision energy was continually altered (3, 10, 20, and 30 eV) during MS acquisition. PCA and OPLS-DA analysis of the generated UPLC–MS data of metabolites extracted from elicited tobacco cells revealed different clustering and different distribution patterns. As expected, ion abundance decreases with increasing collision energy, but, more importantly, results in unique multivariate data patterns from the same samples. Our findings suggest that different collision energy settings should be investigated during MS data acquisition because these can contribute to coverage of a wider range of the metabolome by UPLC–ESI–MS and prevent biased results. Description: Copyright: 2012 Springer-Verlag. This is an ABSTRACT ONLY. The definitive version is published in Analytical and Bioanalytical Chemistry, vol. 404(2), pp 367-372 Wed, 01 Aug 2012 00:00:00 GMT http://hdl.handle.net/10204/6670 2012-08-01T00:00:00Z http://hdl.handle.net/10204/6328 Title: Harvesting sunlight energy: a biophysics approach Authors: Smit, JE; Grobler, AF; Sparrow, RW Abstract: The most efficient light harvesting and energy transfer systems are found in nature as part of the photosynthesis process. In the photosynthetic system light energy is absorbed by antenna chlorophylls and this energy is then passed onto a reaction centre chlorophyll molecule where charge separation occurs in less than 100 ps and at about 95% efficiency. It has been shown that organised connective light harvesting complexes are required for long range energy transfer. By extracting these system fragments and maximising their organisational structure, similar artificial systems for energy sources and transfer system can potentially be developed. As a matrix to stabilize the system we are using a combination of fatty acids and nitrous oxide, rather than conventional phospholipid-based combinations, which enables the production of small, elastic artificial vesicles, called Pheroid™. Previous work has shown that photosynthetic light harvesting material can be incorporated into the Pheroid™. In this study we are characterising the level of organisation through protein aggregation on the incorporated light harvesting systems using absorption spectroscopy. Description: ICWIP 2011: 4th IUPAP International Conference on Women in Physics, Stellenbosch, 5-8 April 2011 Fri, 01 Apr 2011 00:00:00 GMT http://hdl.handle.net/10204/6328 2011-04-01T00:00:00Z