Biology subjects

Dzakovich, M. P.

Publications and source records attributed to Dzakovich, M. P..

7 recordsLinked to original sources

bioRxiv2025

Layers to Leaves: A Suite of Modular 3D Printed Hydroponics Components for Research and Education

Hydroponics is a widely utilized technique to precisely control the plant growing environment and maximize productivity. In some cases, hydroponics systems can be expensive and require specialized expertise to build and maintain. Here, we present a suite of 3D-printed devices that can be constructed into a single or double tower hydroponics system. This system is easily scalable to fit the needs of a user and can be implemented in a variety of research and educational contexts. Components in this suite can be made from inexpensive plastic filament using household-grade 3D printers. The vertical design of these systems allows for users to maximize space-use-efficiency in growth chambers, greenhouses, or even classrooms. We describe the construction of our system and provide example data from a validation study growing spinach (Spinacia oleracea L.) under different salinity conditions. This suite of 3D printed components can be utilized by researchers and educators alike to capitalize on the benefits of hydroponics in a flexible, budget-friendly way.

plant biology↗

bioRxiv2024

A Comprehensive UHPLC-MS/MS and Extraction Method for Spinach (Spinacia oleracea) Flavonoids

Spinach produces an array of unique flavonoids not commonly found in other fruits and vegetables. These molecules likely serve as defense agents against biotic and abiotic stress and may have health beneficial properties for humans. Current methods to analyze spinach flavonoids are incomplete and only capture a portion of this uncharacterized pathway. A comprehensive analysis method is needed to determine how genetics, environmental conditions, and other factors influence spinach flavonoid biosynthesis. We developed and validated a high-throughput extraction and ultra high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method to separate and quantify 39 spinach flavonoid species in 11.5 minutes. Spinach flavonoids without authentic standards were putatively identified using MS/MS fragmentation experiments, precursor scans, and matches to high-resolution MS literature reports. Our extraction method enables up to 48 samples to be extracted in 60 minutes with recovery estimates between 100.5 - 107.8%. To assess the suitability of our method and generate benchmark estimates for 39 spinach flavonoids, we grew a panel of 30 genetically diverse spinach accessions and compared quantification data generated with a traditional or our high-throughput approach. Data generated by either approach were comparable, estimating total flavonoid averages of 75.1 - 170.1 or 93.1 - 187.26 mg/100 g fresh weight for the high-throughput and traditional method, respectively. Many estimates generated by our analysis method represent the first quantitative literature reports of these compounds. These experiments indicate that our extraction and analysis method is efficient, robust, and an important tool needed to study the biosynthesis and biological role of spinach flavonoids.

plant biology↗

bioRxiv2024

Discovery of steroidal alkaloid metabolites and their accumulation in pigs after short-term tomato consumption

ScopeWhole tomato consumption has been shown to be more effective than lycopene alone against chronic disease risks, suggesting other phytochemicals play a role in the health properties of tomato-rich diets. Recently, metabolites of tomato steroidal alkaloids, an understudied class of secondary plant compounds, have been found in plasma, tissues, and urine. However, a comprehensive, targeted analysis to determine which steroidal alkaloid metabolites are present after tomato consumption is lacking. This study profiles and quantifies tomato steroidal alkaloids in blood for the first time. Methods and resultsIn a two-week parallel-arm study, piglets (n = 20) were fed diets containing 10% tomato powder or a macronutrient-matched control. Steroidal alkaloids were extracted from plasma and quantified using LC-MS. Tomatidine and alpha-tomatine were detected in plasma and confirmed with standards, while mass fragmentation spectra aided in identifying 31 additional metabolites representing 9 unique masses. Concentrations averaged to 107.7 nmol/L plasma, comprising of phase I (66%) and phase II (4.5%) metabolites. ConclusionThese results describe the profile and concentration of steroidal alkaloid metabolites in pig plasma after short-term tomato consumption. Our methodology and findings allow for future investigations of tomato steroidal alkaloid bioactivity using physiologically appropriate levels.

pharmacology and toxicology↗

bioRxiv2023

Transcriptomics and Metabolomics Reveal Tomato Consumption Alters Hepatic Xenobiotic Metabolism and Induces Steroidal Alkaloid Metabolite Accumulation in Mice

ScopeTomato consumption is associated with many health benefits including lowered risk for developing certain cancers. It is hypothesized that tomato phytochemicals are transported to the liver and other tissues where they alter gene expression in ways that lead to favorable health outcomes. However, the effects of tomato consumption on mammalian liver gene expression and chemical profile are not well defined. Methods and resultsWe hypothesized that tomato consumption would alter mouse liver transcriptomes and metabolomes compared to a control diet. C57BL/6 mice (n=11-12/group) were fed a macronutrient matched diet containing either 10% red tomato, 10% tangerine tomato, or no tomato powder for 6 weeks after weaning. RNA-Seq followed by gene set enrichment analyses indicated that tomato type and consumption, in general, altered expression of phase I and II xenobiotic metabolism genes. Untargeted metabolomics experiments revealed distinct clustering between control and tomato fed animals. Nineteen molecular formulas (representing 75 chemical features) were identified or tentatively identified as steroidal alkaloids and isomers of their phase I and II metabolites; many of which are reported for the first time in mammals. ConclusionThese data together suggest tomato consumption may impart benefits partly through enhancing detoxification potential.

biochemistry↗

bioRxiv2022

Trait Stacking Simultaneously Enhances Provitamin A Carotenoid and Mineral Bioaccessibility in Biofortified Sorghum bicolor

Vitamin A, iron, and zinc deficiencies are major nutritional inadequacies in sub-Saharan Africa and disproportionately affect women and children. Biotechnology strategies have been tested to individually improve provitamin A carotenoid or mineral content and/or bioaccessibility in staple crops including sorghum (Sorghum bicolor). However, concurrent carotenoid and mineral enhancement has not been thoroughly assessed and antagonism between these chemical classes has been reported. This work evaluated two genetically engineered constructs containing a suite of heterologous genes to increase carotenoid stability and pathway flux, as well as phytase to catabolize phytate and increase mineral bioaccessibility. Kernels from transformed sorghum events were processed into model porridges and evaluated for carotenoid and mineral content as well as bioaccessibility. Transgenic events produced markedly higher amounts of carotenoids (26.4 {micro}g/g) compared to null segregants (4.2 {micro}g/g) and wild-type control (Tx430; 3.7 {micro}g/g). A 200 g serving of porridge made with these transgenic events represents a projected 53.7% of a 4- 8-year-old childs vitamin A estimated average requirement. Phytase activation by pre-steeping flour resulted in significant phytate reduction (9.4 to 4.2 mg/g), altered the profile of inositol phosphate metabolites, and reduced molar ratios of phytate to iron (16.0 to 4.1); and zinc (19.0 to 4.9) in engineered material; suggesting improved mineral bioaccessibility. Improved phytate:mineral ratios did not significantly affect micellarization and bioaccessible provitamin A carotenoids were over 2300% greater in transgenic events compared to corresponding null segregants and wild-type controls. These data suggest that combinatorial approaches to enhance micronutrient content and bioaccessibility are feasible and warrant further assessment in human studies.

plant biology↗

bioRxiv2021

Biosynthesis of Steroidal Alkaloids Are Coordinately Regulated and Differ Among Tomatoes in the Red-Fruited Clade

1.1O_LIWe quantitatively profiled and genotyped two tomato populations representing diversity in the red-fruited clade to address the lack of knowledge regarding the chemical diversity, concentration, and genetic architecture controlling tomato steroidal alkaloids. C_LIO_LIWe grew 107 genetically diverse fresh market, processing, land-race, and wild tomatoes in multiple environments. Nine steroidal alkaloids were quantified using ultra-high performance liquid chromatography tandem mass spectrometry. The diversity panel and a biparental population segregating for high alpha-tomatine, were genotyped to identify and validate quantitative trait loci (QTL) associated with steroidal alkaloids. C_LIO_LILand-races and wild material exhibited higher alkaloid concentrations and more chemical diversity. Average total content of steroidal alkaloids, often dominated by lycoperoside F/G/esculeoside A, ranged from 1.9 to 23.3 mg/100 g fresh weight across accessions. Land-race and wild cherry accessions distinctly clustered based on elevated early or late steroidal alkaloid concentrations. Significant correlations were observed among early and late steroidal alkaloids in a species-dependent manner. A QTL controlling multiple, early steroidal alkaloid pathway intermediates on chromosome 3 was identified by genome wide association (GWAS) and validated in a backcross population. C_LIO_LITomato steroidal alkaloids are diverse in the red-fruited tomato clade and their biosynthesis is regulated in a coordinated manner. C_LI

plant biology↗

bioRxiv2019

A High-Throughput Extraction and Analysis Method for Steroidal Glycoalkaloids in Tomato

Tomato steroidal glycoalkaloids (tSGAs) are a class of cholesterol-derived metabolites uniquely produced by the tomato clade. These compounds provide protection against biotic stress due to their fungicidal and insecticidal properties. Although commonly reported as being anti-nutritional, both in vitro as well as pre-clinical animal studies have indicated that some tSGAs may have a beneficial impact on human health. However, the paucity of quantitative extraction and analysis methods presents a major obstacle for determining the biological and nutritional functions of tSGAs. To address this problem, we developed and validated the first comprehensive extraction and UHPLC-MS/MS quantification method for tSGAs. Our extraction method allows for up to 16 samples to be extracted simultaneously in 20 minutes with 93.0 {+/-} 6.8% and 100.8 {+/-} 13.1% recovery rates for tomatidine and alpha-tomatine, respectively. Our ultra-high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method was able to chromatographically separate analytes derived from 16 tSGAs representing 9 different tSGA masses, as well as two internal standards, in 13 minutes. Tomato steroidal glycoalkaloids that did not have available standards were annotated using high resolution mass spectrometry as well as product ion scans that provided fragmentation data. Lastly, we utilized our method to survey a variety of commonly consumed tomato-based products. Total tSGA concentrations ranged from 0.7 to 3.4 mg/serving and represent some of the first reported tSGA concentrations in tomato-based products. Our validation studies indicate that our method is sensitive, robust, and able to be used for a variety of applications where concentrations of biologically relevant tSGAs need to be quantified.

plant biology↗