Elmer-Rico E. Mojica

Toxicity and reductions in intracellular calcium levels following uptake of a tetracycline antibiotic in Arabidopsis.

Plant responses to natural stresses have been the focus of numerous studies; however less is known about plant responses to artificial (i.e., man-made) stress. Chlortetracycline (CTC) is widely used in agriculture and becomes an environmental contaminant when introduced into soil from manure used as fertilizer. We show here that in the model plant Arabidopsis (Arabidopsis thaliana), root uptake of CTC leads to toxicity, with growth reductions and other effects. Analysis of protein accumulation and in vivo synthesis revealed numerous changes in soluble and membrane-associated proteins in leaves and roots. Many representative proteins associated with different cellular processes and compartments showed little or no change in response to CTC. However, differences in accumulation and synthesis of NAD-malic enzyme in leaves versus roots suggest potential CTC-associated effects on metabolic respiration may vary in different tissues. Fluorescence resonance energy transfer (FRET) analysis indicated reduced levels of intracellular calcium are associated with CTC uptake and toxicity. These findings support a model in which CTC uptake through roots leads to reductions in levels of intracellular calcium due to chelation. In turn, changes in overall patterns and levels of protein synthesis and accumulation due to reduced calcium ultimately lead to growth reductions and other toxicity effects.

Screening of different computational models for the preparation of sol-gel imprinted materials.

Different computational models were used and screened to find a rational way in selecting the appropriate functional silane monomer for the best molecular imprinted xerogel (MIX) formulation. Several functional silane monomers were used and allowed to react with a template model, tetracycline (TC). The resulting template-monomer complex molecules were first optimized and their interaction energies (IEs) were calculated using different computational methods such as semi-empirical methods, ab-initio methods, density functional theory (DFT) methods and solvent model method. The formulations used for calculation were also prepared and their performance in binding with TC was determined using tritium labeled sample. Results showed that the rankings of the different formulations varied with the different computational methods. However, rankings of the IEs of the xerogels are similar to that of the imprinting factor (IF) when HF and B3LYP at SV(P) and SVP basis set levels were used. The best imprinted xerogel, allyltriethoxysilane (AtEOS) ranked first in ten out of the 26 computational models that were screened and at all computational methods at tetramer system.

Determination of the Structures of Molecularly Imprinted Polymers and Xerogels Using an Automated Stochastic Approach

An automated stochastic docking program with a graphical user interface, RANDOMDOCK (RD), has been developed to aid the development of molecularly imprinted polymers and xerogels. RD supports computations with ab initio and semiempirical quantum chemistry programs. The RD algorithms have been tested by searching for the most stable geometries of a varying number of methacrylic acid molecules interacting with nicotinamide. The optimal structures found are either as stable or more stable than those previously proposed for this molecularly imprinted polymer, illustrating that RD is capable of identifying the lowest-energy structures out of a potentially vast number of possible configurations. RD was subsequently applied to determine the most favorable binding sites between silane molecules and tetracycline (TC) as well as TC analogues. Hydrogen bonding between the templates and a silane is an important determinant of stability. Dispersion interactions are also sizable, sometimes dominant, especially between the largest silane and TC analogues not possessing a site readily available for hydrogen bonding. We highlight the importance of exploring the full intermolecular potential energy landscape when studying systems which may not afford highly specific interactions.

Use of computational modeling in preparation and evaluation of surface imprinted xerogels for binding tetracycline

AbstractLinear alkyl alkoxysilanes (methoxy and ethoxy-based) of varying length were used in preparing tetracycline surface imprinted silica xerogels by the sol–gel process. The resulting xerogels were characterized in terms of binding tetracycline (TC) by using tritium-labeled TC. Results showed preferential binding in the ethoxysilane based xerogels in comparison to methoxysilane based xerogels. A computational approach using the interaction energy (IE) between TC and each alkyl alkoxysilane was deduced as a rational way of predicting the formulation that would provide the best analytical performance for a given molecularly imprinted xerogel (MIX). Hartree-Fock calculations revealed an increase in IE as the length of the carbon chain increases until an optimum value at C6 in both alkoxysilanes. This is consistent with the experimental results wherein the C6 xerogel formulation has the highest imprinting factor. These results show the potential of using computational modeling as a rational way of preparing surface imprinted materials. Figure“Use of computational modeling in the preparation and evaluation of surface imprinted xerogels for binding tetracycline”

Formation of N-Ethylmaleimide (NEM)-Glutathione Conjugate and N-Ethylmaleamic Acid Revealed by Mass Spectral Characterization of Intracellular and Extracellular Microbial Metabolites of NEM

ABSTRACT The extracellular and intracellular metabolites formed upon exposure of activated sludge microorganisms to a sublethal concentration of N-ethylmaleimide were monitored by liquid chromatography with ion trap mass spectrometry. The metabolite N-ethylsuccinimido-S-glutathione (m/z 433) was converted rapidly to N-(2-oxoethyl)-2,2-(propionylamino)propanamide (m/z 187) and N-ethylmaleamic acid (m/z 144).

Spectrochemical Behavior of Enzymes Encapsulated within Xerogel

There are many methods that have been used in the encapsulation or immobilization of enzymes. One of them is the sol-gel process, which can be used to produce porous silica materials even at ambient temperature. In this study, several enzymes, namely catalase, lipase, laccase, and glucose oxidase were encapsulated by sol-gel process using tetramethyl orthosilicate (TMOS) as alkoxide. The behavior of the enzymes encapsulated within the gel matrix, or xerogel, was monitored by fluorescence to determine the spectrochemical behavior of the enzymes. The results were compared with the enzymes in solution form. There are minor differences between the spectra in different matrices (xerogel and solution), which can only means that there are conformational changes in enzymes when they were immobilized within the gel matrix.

Raman spectroscopic discrimination of estrogens

Estrogens are a group of steroid compounds found in the human body that are eventually discharged and ultimately end up in sewer effluents. Since these compounds can potentially affect the endocrine system its detection and quantification in sewer water is important. In this study, estrogens such as estrone (E1), estradiol (E2), estriol (E3), and ethynylestradiol (EE2) were discriminated and quantitated using Raman spectroscopy. Simulated Raman spectra were correlated with experimental data to identify unique marker peaks, which proved to be useful in differentiating each estrogen molecules. Among these marker peaks are Raman modes arising from hydroxyl groups of the estrogen molecules in the spectral region 3200-3700 cm-1. Other Raman modes unique to each of the estrogen samples were also identified, including peaks at 1722 cm-1 for E1 and 2109 cm-1 for EE2, which corresponds to their distinctive structures each containing a different set of functional groups. To quantify the components of estrogen mixtures, the intensities of each identifying Raman bands, at 581 cm-1 for E1, 546 cm-1 for E2, 762 cm-1 for E3 and 597 cm-1 for EE2, were compared and normalized against the intensity of a common peak at 783 cm-1. Quantitative analysis yielded most results within an acceptable 20% error.

Conformational differentiation of α‐cyanohydroxycinnamic acid isomers: a Raman spectroscopic study

Two α-cyanohydroxycinnamic acid positional isomers, α-cyano-4-hydroxycinnamic acid (CHCA4) and α-cyano-3-hydroxycinnamic acid (CHCA3), were characterized using Raman spectroscopy. We analyzed the implications of the collected Raman spectral shifts, and verified them through other spectroscopic techniques, to arrive at plausible three dimensional structures of CHCA3 and CHCA4. The positions of these groups were mapped by systematically analyzing the orientation and type of interactions functional groups make in each CHCA isomer. We determined whether or not the carboxylic moieties are forming dimeric links and ascertained the existence of ring-ring π-stacking interactions. We also assessed the nature of the hydrogen bonding between -CN and -OH groups. The results were then taken together to model plausible three dimensional structures for each compound. The data revealed a structure for CHCA4 that matches the published x-ray crystallographic structure. We then applied the same spectral analysis to CHCA3 to reveal its plausible three dimensional structure. The structural details revealed may account for the functional properties of the two α-cyanohydroxycinnamic acid positional isomers.