Zheng-Jiang Zhu

An accelerated workflow for untargeted metabolomics using the METLIN database.

Metabolites, typically recognized as small molecules that are involved in cellular reactions, provide a functional signature of phenotype that is complimentary to the upstream biochemical information obtained from genes, transcripts, and proteins. The high-level of correlation between metabolites and phenotype has created a surge of interest in the field that is reflected in the number of metabolomic publications growing from just a few articles in 1999 to over five thousand in 2011. Although relatively new compared to its genomic and proteomic predecessors, already metabolomics has led to the discovery of biomarkers for disease, fundamental insights into cellular biochemistry, and clues related to disease pathogenesis.1,2

Entrapment of Hydrophobic Drugs in Nanoparticle Monolayers with Efficient Release into Cancer Cells

Gold nanoparticles functionalized with water-soluble zwitterionic ligands form kinetically stable complexes with hydrophobic drugs and dyes. These drugs and dyes are efficiently released into cells, as demonstrated through fluorescence microscopy and cytotoxicity assays. Significantly, there is little or no cellular uptake of particle, making these low toxicity particles promising for delivery applications.

Recognition-mediated activation of therapeutic gold nanoparticles inside living cells

Supramolecular chemistry provides a versatile tool for the organization of molecular systems into functional structures and the actuation of these assemblies for applications through the reversible association between complementary components. Application of this methodology in living systems represents a significant challenge due to the chemical complexity of cellular environments and lack of selectivity of conventional supramolecular interactions. Herein, we present a host-guest system featuring diaminohexane-terminated gold nanoparticles (AuNP-NH2) and complementary cucurbit[7]uril (CB[7]). In this system, threading of CB[7] on the particle surface reduces the cytotoxicity of AuNP-NH2 through sequestration of the particle in endosomes. Intracellular triggering of the therapeutic effect of AuNP-NH2 was then achieved via the administration of 1-adamantylamine (ADA), removing CB[7] from the nanoparticle surface and triggering the endosomal release and concomitant in situ cytotoxicity of AuNP-NH2. This supramolecular strategy for intracellular activation provides a new tool for potential therapeutic applications.

Colorimetric bacteria sensing using a supramolecular enzyme-nanoparticle biosensor.

Rapid and sensitive detection of pathogens is a key requirement for both environmental and clinical settings. We report here a colorimetric enzyme-nanoparticle conjugate system for detection of microbial contamination. In this approach, cationic gold nanoparticles (NPs) featuring quaternary amine headgroups are electrostatically bound to an enzyme [β-galactosidase (β-Gal)], inhibiting enzyme activity. Analyte bacteria bind to the NP, which releases the β-Gal and restores its activity, providing an enzyme-amplified colorimetric readout of the binding event. Using this strategy, we have been able to quantify bacteria at concentrations of 1 × 10(2) bacteria/mL in solution and 1 × 10(4) bacteria/mL in a field-friendly test strip format.

Liquid chromatography quadrupole time-of-flight mass spectrometry characterization of metabolites guided by the METLIN database

Untargeted metabolomics provides a comprehensive platform for identifying metabolites whose levels are altered between two or more populations. By using liquid chromatography quadrupole time-of-flight mass spectrometry (LC-Q-TOF-MS), hundreds to thousands of peaks with a unique m/z ratio and retention time are routinely detected from most biological samples in an untargeted profiling experiment. Each peak, termed a metabolomic feature, can be characterized on the basis of its accurate mass, retention time and tandem mass spectral fragmentation pattern. Here a seven-step protocol is suggested for such a characterization by using the METLIN metabolite database. The protocol starts from untargeted metabolomic LC-Q-TOF-MS data that have been analyzed with the bioinformatics program XCMS, and it describes a strategy for selecting interesting features as well as performing subsequent targeted tandem MS. The seven steps described will require 2–4 h to complete per feature, depending on the compound.

The Role of Surface Functionality on Acute Cytotoxicity, ROS Generation and DNA Damage by Cationic Gold Nanoparticles

Gold nanoparticles (AuNPs) are promising materials for biomedical applications [1,2] due to their tunable surface properties [3] and extraordinary stability.[4] Additionally, the inert core material reduces the potential for toxicity issues arising from particle degradation.[5] The size regime and concomitant geometrical outcomes including high degree of curvature, however, generates the potential for toxicity.[6,7] Generally, the toxicity of AuNPs depends on size, shape, the degree to which they aggregate, and their surface properties[8,9] Recently, several studies on the short-term cytotoxicity of AuNPs[10] and quantum dots[11] have focused on size,[12,13] shape,[14,15] and charge.[16] To date, however, issues such as ligand hydrophobicity have not been systematically explored.

Intracellular Delivery of a Membrane-Impermeable Enzyme in Active Form using Functionalized Gold Nanoparticles

Gold nanoparticles were coated with a short peptide to promote intracellular delivery of membrane-impermeable proteins. Through microscopy and enzyme assays, we demonstrated the particles were able to transport functional enzymes into a variety of cell lines. Significantly, the transported proteins were able to escape from endosomes. Moreover, these particles showed no apparent cytotoxicity.

Direct delivery of functional proteins and enzymes to the cytosol using nanoparticle-stabilized nanocapsules.

Intracellular protein delivery is an important tool for both therapeutic and fundamental applications. Effective protein delivery faces two major challenges: efficient cellular uptake and avoiding endosomal sequestration. We report here a general strategy for direct delivery of functional proteins to the cytosol using nanoparticle-stabilized capsules (NPSCs). These NPSCs are formed and stabilized through supramolecular interactions between the nanoparticle, the protein cargo, and the fatty acid capsule interior. The NPSCs are ~130 nm in diameter and feature low toxicity and excellent stability in serum. The effectiveness of these NPSCs as therapeutic protein carriers was demonstrated through the delivery of fully functional caspase-3 to HeLa cells with concomitant apoptosis. Analogous delivery of green fluorescent protein (GFP) confirmed cytosolic delivery as well as intracellular targeting of the delivered protein, demonstrating the utility of the system for both therapeutic and imaging applications.

Stability of quantum dots in live cells.

Quantum dots (QDs) are highly fluorescent and photostable, making them excellent tools for imaging. When using these QDs in cells and animals, however, intracellular biothiols (e.g., glutathione and cysteine) can degrade the QD monolayer compromising function. Here, we describe a label-free method to quantify the intracellular stability of monolayers on QD surfaces that couples laser desorption/ionization mass spectrometry (LDI-MS) with inductively coupled plasma mass spectrometry (ICP-MS). Using this new approach we have demonstrated that QD monolayer stability is correlated with both QD particle size and monolayer structure, with proper choice of both particle size and ligand structure required for intracellular stability.

Effect of surface charge on the uptake and distribution of gold nanoparticles in four plant species.

Small (6−10 nm) functionalized gold nanoparticles (AuNPs) featuring different, well-defined surface charges were used to probe the uptake and distribution of nanomaterials in terrestrial plants, including rice, radish, pumpkin, and perennial ryegrass. Exposure of the AuNPs to plant seedlings under hydroponic conditions for a 5-day period was investigated. Results from these studies indicate that AuNP uptake and distribution depend on both nanoparticle surface charge and plant species. The experiments show that positively charged AuNPs are most readily taken up by plant roots, while negatively charged AuNPs are most efficiently translocated into plant shoots (including stems and leaves) from the roots. Radish and ryegrass roots generally accumulated higher amounts of the AuNPs (14−900 ng/mg) than rice and pumpkin roots (7−59 ng/mg). Each of the AuNPs used in this study were found to accumulate to statistically significant extents in rice shoots (1.1−2.9 ng/mg), while none of the AuNPs accumulated in the shoots of radishes and pumpkins.