Nutrient management plays a critical role in sustaining high-rate biological hydrogen production, whether through photobiological or dark-fermentation systems. The availability of nitrogen, phosphorus, and trace metals directly influences microbial metabolism, hydrogenase enzyme activity, and overall system stability. Without proper nutrient balance, microbial consortia experience reduced efficiency, leading to lower hydrogen yields and potential process failure.

In photobiological hydrogen production, cyanobacteria and microalgae rely on nitrogen and phosphorus for growth and photosynthetic activity. Nitrogen is a key component of chlorophyll, proteins, and hydrogenase enzymes, while phosphorus is essential for ATP synthesis and nucleic acid formation. Trace metals such as iron, nickel, and magnesium serve as cofactors for hydrogenases and nitrogenases. For example, iron is a central component of [FeFe]-hydrogenases, which catalyze proton reduction in many green algae. Nitrogen limitation often triggers a metabolic shift from biomass growth to hydrogen production in certain cyanobacteria, as seen in strains of *Anabaena* and *Nostoc*, where nitrogenase activity increases under nitrogen-deprived conditions. However, prolonged nitrogen starvation can lead to cell deterioration and reduced hydrogen output.

Dark-fermentation systems, which utilize anaerobic bacteria like *Clostridium* and *Enterobacter*, also depend heavily on nutrient availability. Nitrogen is required for protein synthesis and enzyme production, while phosphorus supports energy metabolism. Trace metals such as iron, nickel, and selenium are critical for hydrogenase and ferredoxin function. For instance, *Clostridium acetobutylicum* exhibits optimal hydrogen production when iron concentrations are maintained within a narrow range, as both deficiency and excess impair hydrogenase activity. Similarly, nickel is a cofactor for [NiFe]-hydrogenases in many fermentative bacteria, and its absence can drastically reduce hydrogen yields.

Nutrient limitations disrupt microbial activity in predictable ways. Nitrogen scarcity slows cell division and enzyme synthesis, leading to prolonged lag phases and reduced hydrogen productivity. Phosphorus deficiency impairs energy metabolism, limiting ATP availability for hydrogen-producing pathways. Trace metal shortages directly inhibit hydrogenase and nitrogenase function, while excessive concentrations can cause toxicity. For example, high iron levels may generate reactive oxygen species, damaging cellular components and reducing microbial viability.

Recycling spent media is a promising strategy to reduce nutrient input costs and improve sustainability. In photobiological systems, nutrient recovery can be achieved through biomass harvesting and subsequent extraction. Microalgal biomass, after hydrogen production, retains significant amounts of nitrogen and phosphorus, which can be recovered via anaerobic digestion or thermal treatment. The resulting digestate or ash can be reintroduced into fresh growth media, closing the nutrient loop. Some studies have demonstrated successful media recycling with *Chlamydomonas reinhardtii*, where up to 70% of the original nutrient content was reused without significant yield loss.

Dark-fermentation systems also benefit from spent media recycling. Effluent from hydrogen production contains residual volatile fatty acids, ammonia, and phosphate, which can support subsequent fermentation batches. However, inhibitors like organic acids and residual solvents may accumulate over multiple cycles, necessitating periodic treatment. Techniques such as membrane filtration, electrodialysis, or adsorption can remove inhibitory compounds while retaining essential nutrients. For example, *Thermoanaerobacterium thermosaccharolyticum* cultures have been maintained for multiple batches using recycled media supplemented with only trace metals, reducing overall nutrient demand by 40-50%.

A comparative analysis of nutrient requirements in photobiological and dark-fermentation systems reveals key differences. Photobiological systems generally demand higher nitrogen inputs due to the photosynthetic apparatus requirements, while dark-fermentation systems prioritize trace metals for enzymatic activity. Phosphorus needs are similar in both systems but must be carefully balanced to avoid precipitation or lock-up in insoluble forms.

Optimizing nutrient management involves continuous monitoring and adaptive supplementation. Real-time sensors for nitrogen, phosphorus, and trace metals can help maintain optimal concentrations, preventing both limitation and toxicity. Automated dosing systems have been successfully integrated into pilot-scale photobioreactors and fermenters, ensuring stable hydrogen production over extended periods.

The future of nutrient management in biological hydrogen production lies in integrated systems that combine nutrient recycling with waste valorization. Coupling photobiological and dark-fermentation processes could allow cross-utilization of effluents, where nitrogen-rich effluent from dark fermentation supports algal growth, while algal biomass provides micronutrients for fermentative bacteria. Such synergies could significantly reduce external nutrient demands while enhancing overall hydrogen output.

In summary, nutrient management is a cornerstone of efficient and sustainable biological hydrogen production. Proper balancing of nitrogen, phosphorus, and trace metals maximizes microbial activity and hydrogen yields, while recycling strategies minimize resource consumption. Advances in nutrient recovery and system integration will further enhance the viability of large-scale biohydrogen systems.