Microgreens for Pea(ce) - Zinc Biofortification through Seed Nutripriming to Fight Hidden Hunger

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Background

Hidden hunger is a form of malnutrition where individuals, despite having a sufficient calorie intake, lack the essential vitamins and minerals required for proper growth and immune functioning. This deficit of micronutrients is not always visible, but leads to health consequences for all vulnerable populations. A study conducted by two nutritional scientists at UC Davis found that overall, “17.3% of the world’s population” has been identified as at risk for a zinc deficiency (Wessells & Brown, 2012). This number is representative of over one billion individuals, and additional data based on a nutrient composition database found that these deficiencies are most prevalent in “South and Southeast Asia, Sub-Saharan Africa and Central America” (Wessells & Brown, 2012). There is a critical need to implement more accessible and sustainable nutritional options in these regions. Additionally, according to a 2025 joint report from the Food and Agriculture Organization (FAO) and the World Food Programme (WFP), an “estimated 31.9 percent of people (2.60 billion)” cannot afford a healthy diet, contributing to malnutrition due to mineral deficiencies (FAO et al., 2025).

Microgreens are a highly promising intervention method, since they are nutrient-dense plants that can be grown using low-cost methods, like hydroponics or growing trays. Since they require low space and water resources, these plants are ideal for a variety of applications, such as combating malnutrition and supplementing daily diets (Seth et al., 2025). However, standard microgreens still lack sufficient concentrations of minerals, such as zinc, to overcome acute deficiencies. This project focuses on a technique to increase the density of micronutrients in the crop, called agronomic biofortification. This method is comparable to genetic engineering solutions and fertilizer applications, but is more directly applicable to marginalized communities because it is easily scalable. Unlike zinc supplements, biofortified microgreens provide a more natural and sustainable food source.

Literature Review

Seed nutripriming, the process of soaking microgreen seeds in a mineral solution, has been previously identified as a practical strategy for agronomic biofortification that relies on the plant’s ability to absorb nutrients during the initial imbibition phase. An analysis by researchers at the University of Calicut confirmed that, at certain concentrations, zinc priming successfully enhanced the germination percentage and seed vigor of crops (Veena & Puthur, 2021). These are important physiological markers used to prove plant biochemical health, but it also introduces the concept of a toxicity threshold. Veena and Puthur documented that extremely high priming concentrations can induce oxidative stress and decrease positive outcomes. This is why precise testing should be completed across many concentrations to find the exact, optimal balance between nutritional benefits and crop health. That study reinforces the premise of this experiment, which focuses on optimization and the eventual real-world implementation of these findings.

The direct application of nutripriming to microgreens has provided the foundation for this current research. A 2023 experimental study by Pennsylvania State University directly analyzed the effects of zinc sources and concentration levels on the nutritional content of both sunflower and pea microgreens (Poudel et al., 2023). Their results confirmed that zinc sulfate supplementation at a 200 ppm concentration was highly effective for the delicate plants. Additionally, the treatment produced more plant biomass without negatively impacting the early growth rates or germination (Poudel et al., 2023). It directly compares the effectiveness of zinc sulfate and zinc oxide in enhancing nutrient levels, and the positive findings demonstrate that this is a viable solution for biofortifying microgreen plants. The areas that still remain under-investigated involve adapting for settings with limited resources.

The majority of current biofortification literature, such as work by Dr. Pradip Poudel, utilizes a soil or peat-perlite substrate mix as a physical buffer to grow the microgreen plants. The goal is to modify these experiments to not depend on a growing medium and create a novel project tailored towards displaced communities globally. It is necessary to review biofortification studies conducted using soilless growing mediums to address the methodological gap that will then be applied to the pea microgreens. A paper focusing on Brassicaceae microgreens found experimental data on whether the plant could be biofortified without causing significant growth restrictions (Di Gioia et al., 2019). This is a large concern when using a soilless growing method that provides no supplemental nutrients. The scientists reviewed the mineral treatments and quantified tissue material content from their trials, which were determined to have positive benefits on growth and nutritional metrics as a result from the zinc sulfate and iron treatments (Di Gioia et al., 2019). This proves that nutripriming techniques are feasible in a soilless framework and appropriate to use for further applications.

Furthermore, research conducted by Gunjal et al. (2024) confirms that microgreens grown in soilless media can maintain high levels of bioactive compounds and antioxidant properties. However, these biochemical markers were highly sensitive to the tested nutrient concentrations. This proves that morphological parameters, such as shoot and root development, are directly influenced by the mineral availability in soilless systems (Gunjal et al., 2024). Nutripriming under these unique conditions requires more precise concentration management to avoid any unintended physiological plant stress. 

Finally, controlling external factors that also interact with the experiment was crucial to obtaining accurate results. For example, a peer-reviewed paper on the relationship between light intensity and zinc biofortification levels analyzed how this variable impacted the plant metabolic profile (Poudel et al., 2025). It specifically found that varying light levels increased the synthesis of beneficial compounds, like flavonoids and phenolic acids, in microgreens. Since the conclusion confirms that nutrient enrichment is correlated to specific environmental conditions, they must be held constant during experimentation so only one variable is studied at a time. This paper was the basis for the constraints that were standardized, such as light exposure, room temperature, and humidity.

Gap Statement

While existing studies have focused on studying zinc nutripriming within controlled environments using soil and other physical mediums, this project uses soilless microgreen cultivation conditions without any traditional grow medium to better stimulate conflict environments where soil and infrastructure are unavailable (Poudel et al., 2023). Experiments have confirmed that this has the potential to increase the biomass and zinc content of plants, but it has never been applied to Arvika pea seeds and optimized to the toxicity boundary (Di Gioia et al., 2019). Also, this addresses the feasibility of using microgreen growing kits within humanitarian aid packages to bridge the gap between research and actual implementation. This approach considers the limitations in adaptability and measurement techniques to find a solution for hidden hunger in complex environments that has never been done before.

The project uses an experimental method to directly collect data in regards to this topic. The overall research question is: How does seed nutripriming with zinc sulfate at concentrations ranging from 0 to 250 ppm influence the germination rate, shoot and root development, and zinc bioavailability of pea microgreens grown using a soilless medium? The aim is to identify the optimal zinc sulfate concentration for maximizing bioavailable zinc without causing toxicity and inferring the link between nutripriming and the reduction of phytic acid levels in the seeds. Phytic acid is a molecule in seeds that binds to zinc, and other mineral cations, to form insoluble compounds which the human body cannot easily absorb (Kumar et al., 2021). High consumption of phytic acid can reduce bioavailability of the nutrients, so limiting it is an important factor in improving nutritional outcomes.

Hypothesis

The hypothesis is that if speckled pea seeds are nutriprimed with increasing concentrations of zinc sulfate, then the resulting microgreens will exhibit enhanced development characteristics, specifically through increased shoot height, root length, and biomass after harvest when compared to the control, until reaching the phytotoxic threshold after 50 ppm. This study utilizes physical growth markers as indicators of nutrient uptake, since optimal zinc levels have been found to reliably increase plant vigor by catalyzing the enzymatic process (Veena & Puthur, 2021). Due to the lack of laboratory access, any changes in microgreen growth and health statistics are reasonably inferred to directly reflect the change in zinc sulfate concentration levels. Though the precise phytic acid levels cannot be quantified using the PA:Zn molar ratio without advanced equipment, tracking changes in the biomass is a practical measurement of biofortification success that can be applied to any environment. This topic covers the broader fields of plant physiology and nutritional science and aligns with the United Nations Sustainable Development Goal 2, which aims to end all forms of malnutrition and hunger worldwide by 2030 (FAO et al., 2025). This further explains why this study is so pivotal because of its practical and global applications to address food insecurity.

Experimental Design and Methodology

The main subjects of this experiment were speckled pea microgreen seeds biofortified with different concentrations of zinc sulfate. The scientific name of the general pea species is Pisum sativum, and it includes the main subgroups of English peas, snow peas, and sugar snap peas (True Leaf Market, 2026). The Arvika pea seed was specifically selected for use in this study. This type is a popular variety for growing microgreens and cover crops because of its reported high germination rates and nutritional value relative to other forage peas. This factor, alongside its widespread availability, made them an ideal choice for this zinc biofortification study aimed at addressing nutritional deficiencies.

As part of the selection process, the Arvika pea microgreen seeds were randomly chosen and separated according to an established sampling plan. The seeds were acquired from True Leaf Market in 1 lb seed packages, containing approximately 2,300 seeds each (True Leaf Market, 2026). While they were not handpicked for specific size, a manual quality control check was performed and it was ensured that all of the seeds were fully developed and had an intact seed coat, known as the testa. This ensured that any germination failure was due to the experimental treatments rather than prior seed damage. 

The methodology used for this research was an experimental design. A total of 2,448 microgreen seeds were grown, with 204 seeds assigned to each of the four treatment conditions and then duplicated over three trials for scientific accuracy. This value was calculated by filling one entire microgreen growing tray with the speckled pea seeds, a total of 340 seeds. An online microgreen seed density calculator, developed by Dr. Francesco Di Gioia, provided valuable information on the ideal seed weight (g) according to the specific dimensions of the growing tray (Di Gioia, n.d.). These factors determined that, in this experiment, only 60% of each tray was to be filled. This calculated value was 204 seeds, chosen to ensure that they can be individually separated to record germination observations and that the microgreens would not be overly crowded by the end of the experiment. The overall design of this experiment was structured around the following sampling plan, which showcases each condition tested:

The primary data in this experiment was collected directly using the established design process. Minimal secondary data was relied on during this process and this established methodology ensures that the plant density setup is reproducible across all trials. 

Materials

The materials for this experiment were chosen and utilized in accordance with previously established botanical research standards. Alongside the speckled pea microgreen seeds, zinc sulfate monohydrate (ZnSO₄·H₂O) from the brand Pacific Pelican was used as the biofortification agent. The purity and industrial-grade level of this specific compound was critical to the experiment’s accuracy. If the product were not high-purity, the actual amount of elemental zinc would be below the calculated stoichiometric value and skew the parts per million (ppm) concentrations.

The preparation of the nutrient solutions required 1 L of distilled water to make the stock solution without contamination. Additionally, 12 plastic microgreen trays were purchased to grow the seeds in a consistent environment across all trials. To monitor the conditions of the growing trays, a digital thermometer and hygrometer was used to collect precise temperature and humidity data. These internal and external values were kept constant to validate that any changes were the result of zinc concentrations rather than environmental stress. For data collection, a camera was used to take pictures of the microgreen plants at every stage of the testing process, in 12 hour intervals when additional data measurements and observations were recorded.

Finally, there was precision in the data collection process through using advanced image analysis tools, more specifically, the National Institute of Health’s (NIH) ImageJ software (Schneider et al., 2012) and the Canopeo Leaf Area Calculator (Patrignani & Ochsner, 2015). These tools have the ability to measure plant development with a higher level of accuracy than is possible manually. Still, to confirm the reliability of these tools, a simple ruler was used for the calibration setup and also basic growth measurements. 

Procedure

The procedure for this experiment was adapted from previously conducted studies to simulate a resource constrained and soilless environment. The main point of reference was a Pennsylvania State University research article, which is part of a project titled “Food Resilience in the Face of Catastrophic Global Events” that explores the broader field of study in depth (Pennsylvania State University, 2025). This experiment monitors pea microgreen shoot height, radicle emergence, root length, root mat thickness, canopy density, and fresh weight at harvest, as indicators for zinc bioavailability. These factors were based on how micronutrient uptake can lead to quantifiable increases in plant development without the need to use spectroscopic equipment. Zinc sulfate monohydrate was tested at 0, 50, 200 and 250 ppm concentration levels within the microgreens.

Before initiating the procedure, the stoichiometric yield of elemental zinc from the Pacific Pelican zinc sulfate monohydrate (ZnSO₄·H₂O) was calculated for precision. Given the molar mass of 179.456 g/mol and an elemental zinc mass of 65.38 g/mol, the product was determined to contain 36.43% of zinc after dividing the values. This was applied in determining the amount required for each concentrated solution. A digital milligram scale was used to measure 0.137 g of the product for the 50 ppm solution, 0.549 g for the 200 ppm solution, and 0.686 g for the 250 ppm solution. Each was dissolved into exactly 1 L of distilled water to create a stock solution distributed equally across all three trials for each concentration.

In the nutripriming phase, 12 groups of 204 Arvika pea seeds were submerged in 300 mL of their assigned solutions for a 12 hour imbibition period, with de-ionized water for the control group. The temperature and humidity of the environment was kept constant to ensure that there were no unintended variables being tested. Afterwards, the seeds were evenly distributed into their respective plastic microgreen trays, according to the sampling plan, ensuring that none overlapped. They were then sprayed with de-ionized water and placed in a dark environment (under a cardboard box) for 48 hours to simulate germination phase growth. Taking careful observations in this phase was very important for calculating the germination rate of the seeds for each concentration. After the two day period, the trays were transferred into a lighter environment where the temperature ranged from 20 to 22°C and humidity levels were considerably lower.

The data collection occurred every 12 subsequent hours over the remaining 10 day testing period. Qualitative observations were taken as the plants were monitored for visual signs of phytotoxicity like chlorosis (leaf yellowing) or stunted growth. The growth metrics were further analyzed through NIH’s ImageJ tool (Schneider et al., 2012) and the Canopeo Leaf Area Calculator app (Patrignani & Ochsner, 2015) for more precise measurements for root length and canopy density. The last step was harvesting the microgreens at the end of the testing period by cutting the shoots 1 cm above the tray line.

Results

The above data table summarizes the growth of all tested pea microgreen plants across the three experimental trials for all concentrations: 0, 50, 200, and 250 ppm. The first part of the growth phase was focused on observing germination rate, measured on Day 2 when the radicle emerged from the seed and was at least one cm long for each treatment. The pea seeds germinated consistently at a rate of over 95% for all four zinc concentration levels. The control group of 0 ppm was used to indicate the baseline throughout all testing variables. In this case, the highest germination rate 99.02% was observed at the 50 ppm concentration, which was a 0.74% increase over the control. However, there was a downward trend for the concentrations above 50 ppm, with the 250 ppm group decreasing 1.86% from the baseline. The overall germination success for all trials was important since the seedlings had to thrive in order to be further examined throughout the growth period.

After the germination phase ended, shoot height was used as one of the primary factors to observe microgreen growth. The control group had an average shoot height of 23.5 cm by the end of the experiment on Day 10. The 50 ppm group had an average height of 24.67 cm, again ranking over the baseline by 4.98%. The inhibitory effects of the higher zinc concentrations started at 200 ppm, where shoot height decreased to 22.53 cm. It was the most pronounced at 250 ppm when the average height was 22% lower relative to the control group, and only reached an average of 18.33 cm.

The average root length of the microgreens showed the most statistical variation across all zinc sulfate concentrations. Each value recorded was confirmed using the NIH’s ImageJ root tracing software (Schneider et al., 2012). The 50 ppm group resulted in an average root length of 12.30 cm, which was an overall 24.62% increase over the control group’s 9.87 cm. This was the largest positive percent change recorded across all of the growth metrics in this study. Conversely, the root length for the 250 ppm concentration plants decreased 62.21% from the baseline. This reduction was also significantly more than any of the other dependent variables tested.

In addition to the primary root lengths, the overall thickness of the root mat was taken to find the density of the roots within each growing tray. Following the previous observed data trends, the 50 ppm group reached a thickness of 3.2 cm or 18.52% over the control group. Even though root length decreased at 200 ppm, the root mat thickness was slightly above the baseline at 2.9 cm. This was unexpected because even though the individual main roots were shorter for this testing condition, the entire tray had a greater density. At 250 ppm, the thickness decreased by 33.33% from the baseline as expected.

Canopy density was measured and quantified using the Canopeo image analysis tool to estimate the percent of surface area covered by the microgreen foliage (Patrignani & Ochsner, 2015). The baseline had a density of 42.58% and the highest density canopy average was 45.72% by the 50 ppm group. The 200 and 250 ppm tests had a thinner canopy up to 24.93% below the control. The overall trends of lower values for these two testing conditions led to shorter stems and leaves, which resulted in a reduced canopy density.

Finally, the fresh weight of the plants at harvest decreased as the zinc sulfate concentrations increased, with the exception of the 50 ppm condition that had the maximum 121.75 g yield. This is consistent with inferred results based on the other quantitative and qualitative factors over the ten day time period. For the last two days of the growth cycle, the qualitative observations showed slower growth overall across all dependent variables, which indicated the end of the testing phase. 

Another notable observation made during this experiment was the changing water color during the imbibition phase. As the seeds were soaking in their respective zinc sulfate solutions, with concentrations of 0, 50, 200, and 250 ppm, prior to germination, the water clarity changed from clear to purple, with a deeper color being observed for the higher concentration trials.

Additionally, there was a qualitative difference in microgreen hardiness across all four testing concentrations. The plants in the 0 to 200 ppm groups consistently grew straight for the entire ten day period, but the 250 ppm trials were noticeably thinner and less rigid by day 7. These observations are supported by the quantitative data that was collected, because the 250 ppm group performed consistently worse for all measured dependent variables.

Discussion 

The quantitative and qualitative results of this experiment indicated that 50 ppm of zinc sulfate monohydrate is the optimal concentration for nutripriming the speckled pea microgreen seeds. This concentration maximized the development and final biomass output of the plant without crossing the threshold into phytotoxicity. Even though the baseline, 0 ppm of zinc sulfate, produced healthy microgreens the 50 ppm treatment group had significantly better results across all of the measured dependent variables and visual observations. For example, the average fresh harvest weight reached an average of 121.75 g, which was a 15.31% increase from the 105.58 g control plants. The percent difference from the control was also consistently more positive than the other tested concentrations. This increase is because zinc acts as a catalyst for protein synthesis, which is how the 50 ppm concentration maximized enzymatic activity through absorption during the 12 hour imbibition period.

This study supports general pre-existing academic literature concluding that zinc is one of many nutrients that can be used to enhance the vigor of microgreen plants. However, there is a discrepancy between these results and research conducted by Pennsylvania State University, which identified 200 ppm as the most effective for increasing zinc sulfate uptake (Poudel et al., 2023). In this experiment, the 200 and 250 ppm concentration groups exhibited clear signs of growth decline. In the average measurements of the 250 ppm test, the root length and canopy density both dropped by 62.21% and 24.93% respectively. This study is working to fill informational gaps by proving that in a soilless growing environment, speckled pea seeds have a lower toxic threshold than previously believed. The observed purple discoloration of the water during the soaking period provides further evidence for the threshold. The color change was likely due to the leaching of anthocyanins, which are red, purple, or blue pigments within the plant tissues produced to combat oxidative stress (Rawat et al., 2025). In the higher concentration trials, this was a distinct visual indicator of the inhibited metabolic activation as germination occurred.

Even despite the observed statistical and visual trends in the data, there were some limitations in concluding definite results from this experiment. The biggest limitation was the potential for oxygen deprivation, known as hypoxia, during the imbibition phase. The seeds were submerged in still deionized water, so the higher concentrations of zinc intensified their lack of oxygen and possibly led to the stunted growth metrics observed in the 200 and 250 ppm groups. This explains why these results differed slightly from other published research studies, since without aeration the plants could not metabolize the higher levels of zinc. Additionally, no direct tissue analysis was conducted to quantify the amount of zinc that was transferred into the shoots and leaves of the microgreen plant. This was due to a lack of access to atomic absorption spectroscopy as a measurement method because of cost. Without chemical verification of the plant matter this experiment is unable to determine how much zinc would actually be transferred to the human consumer.

Conclusion

The overall findings from this study establish that the 50 ppm zinc sulfate monohydrate concentration was optimal, according to analysis of all tested dependent variables, and was the threshold for toxicity in the experiment. Beyond this point, the data suggested that the benefits of biofortification are hindered by physiological stress placed on the microgreen plant. This has created a boundary and beneficial framework for the applications of nutripriming in future food security initiatives.

The real-world and global impact of this study is that nutripriming with pea microgreen seeds has the potential to turn a simple crop into a portable and high-density nutrient option for many individuals currently displaced by climate disaster or war. This directly addresses the “hidden hunger” food crisis, where the lack of consumption of essential micronutrients stunt human growth and development. Unlike other synthetic supplements utilized to address this concern, these nutritious microgreens can provide the added benefits of fiber and vitamins. They are also a self-reliant method for maintaining health in low-resource environments because microgreens require limited space and water, and can be grown in a short period of time. In the broader fields of plant physiology and nutritional science, this experimental study highlights the gaps in how researchers understand nutrient uptake in soilless nutripriming systems. The optimal threshold that was found at 50 ppm is also essential for understanding further applications of these findings in humanitarian aid. It also prevents the accidental implementation of zinc concentrations that could lead to crop failure in harsher environmental settings.

Future Research

To bridge the gap between this research and actual implementation, a sample humanitarian growing kit was 3D printed as a tangible prototype of this study’s applications. This kit is contained inside the created soilless growing tray, and includes a pack of Arvika pea seeds, a spray bottle and standardized measuring tools. Its purpose is to allow individuals to grow their own plants for consumption in environments where agricultural infrastructure is unavailable.

The experimental findings, alongside the created aid kit, show how nutripriming can fight hidden hunger by improving the quality and nutritional content of microgreens. Microgreens are a simple and efficient food source because they require minimal water and resources to thrive, while delivering important nutrients into the body. This research also provides expected and scalable results for organizations like the United Nations World Food Program (WFP) to implement into their future initiatives.

In order to address the limitations of this specific study, researchers should implement an aerated soaking protocol, using tools such as an aquarium air pump, to maintain dissolved oxygen levels. This would prevent the potential for hypoxia and clarify whether the growth decline at 200 ppm was due to zinc toxicity or a lack of oxygen due to the experimental procedure used. Additionally, future studies must have chemical verification of the plant tissue to determine the bioavailable zinc percentage transferred to the consumer. As mentioned previously, the most effective method would be the use of atomic absorption spectroscopy to determine the zinc sulfate content within the shoots specifically. Furthermore, the use of a timelapse or more consistent photography over a 6 hour timespan could reduce human error in measuring the radicle emergence. This procedure change could result in a more precise timeline for the overall metabolic activity accelerated through the process of zinc nutripriming.

Finally, to transition from this theoretical experiment into a distributable humanitarian kit, the next steps for researchers should center around more logistical and environmental variables. For example, more studies should be conducted on how pre-primed and re-dried pea microgreen seeds perform after 3 to 6 months of storage. It is vital to ensure that these seeds can still maintain an 80% or greater germination rate after being subjected to uncontrollable conditions, such as fluctuating temperature and humidity. There is also a need for the development of simplified mixing tools or procedures so that families can create the 50 ppm solution using improvised containers. One idea that could be expanded on is the potential for pre-measured zinc packets within a larger microgreen growing kit. Lastly, the impact of water quality should be tested for this application, because high salinity or unique pathogens could inhibit the uptake of zinc. These are some of the final steps needed to confirm that this nutripriming method could be reliably utilized to improve the quality of life for individuals across the globe.