The simultaneous application of fulvic acid and protein hydrolysate biostimulants enhances cucumber responses to Fe deficiency

Iron (Fe) is widely recognized as a critical factor in limiting crop production; however, eco-friendly strategies to address its deficiency are still required. The use of biostimulants has displayed promising results in mitigating Fe deficiency. Our hypothesis was that the combined application of two biostimulants with distinct molecular structures - fulvic acid (FA) and protein hydrolysate (PH) - could be more effective than the use of a single compound. The simultaneous presence of FA and PH (MIX) in a Fe-free nutrient solution led to a redistribution of endogenous Fe, resulting in a higher leaf SPAD index. Furthermore, the addition of FeCl₃ as a Fe source (resupply) in MIX-treated plants enhanced the biostimulant effect, as evidenced by increased dry root and shoot weight and a more developed root system. In addition, the expression of Strategy-I-related genes, CsFRO1 and CsIRT1, remained elevated. These effects can be attributed to improved interaction between the roots and biostimulants through the formation of the FA-PH complex, as demonstrated by circular dichroism and isothermal titration calorimetry analyses.