Soybean [Glycine max L. (Merr.)] and maize (Zea mays L.) are some of the most important crops worldwide (Xu et al., 2020), with global areas of approximately 120 and 197 million hectares, respectively, in 2019 (Food and Agriculture Organization of the United Nations, 2019). The increasing demand for food has heightened interest in technologies for optimizing agricultural production without increasing the consumption of natural resources (Schröder et al., 2019). One such strategy is foliar fertilization, which is an important technique for additional nutrient supply, increasing plant growth and production, even when plants are not nutrient deficient (Rosolem, 2002). Because of the small amounts of Mo required by most plants, Mo application with solid fertilizers at seeding is not always done; therefore, the supply of Mo to field crops may be neglected. The application of small doses of nutrients strategically at specific phenological stages can stimulate carbon metabolism and enhance tolerance to abiotic stresses such as drought stress (Primavesi, 1978; Alexander, 1986; Hussain et al., 2021; Rodrigues et al., 2021). However, gaps remain in the understanding of the effects of supplemental fertilization with micronutrients, especially molybdenum (Mo), on plant metabolism and production under field conditions.

Mo is a component of enzymes that are essential for the absorption, assimilation and transport of nitrogen (N) in plants (Mendel, 2013; Bittner, 2014). Consequently, Mo deficiency or supplementation indirectly affects the products of N metabolism (Marschner, 2012). Mo is typically applied with the aim of stimulating biological nitrogen fixation (BNF) and nitrate reduction (Calonego et al., 2010; Mercante et al., 2011; Crusciol et al., 2019). Under Brazilian field conditions, the application of Mo via seed or leaves has been shown to be efficient in increasing nodulation, nitrate reductase activity, protein content, leaf area, number of pods, and grain yield (Calonego et al., 2010; Almeida et al., 2013; Silva et al., 2017; Crusciol et al., 2019); however, recent research has demonstrated that Mo also directly impacts photosynthesis due to its involvement in chlorophyll biosynthesis and stability of the photosynthetic apparatus (Yu et al., 2006; Imran et al., 2019). In addition, enhancing photosynthesis by adding Mo increases water use efficiency (WUE), leading to greater tolerance of abiotic stresses (Wu et al., 2020). However, these results were obtained in experiments performed under controlled conditions, and the actual effects of Mo supplementation on crop physiology and productivity under field conditions remain unclear. We hypothesized that Mo stimulates N and carbon metabolism to increase photosynthesis and plant productivity. To test this hypothesis, we investigated the effect of foliar application of Mo on the physiology and grain yield of soybean and maize.

Mo plays an important role in several plant metabolic processes, most notably biological nitrogen fixation and nitrate reduction (Calonego et al., 2010; Steiner and Zoz, 2015; Crusciol et al., 2019). Recent studies have reported direct effects of Mo on photosynthesis via metabolic pathways that are not yet fully understood (Yu et al., 2006; Imran et al., 2019; Rana et al., 2020). However, most of these works were performed under controlled conditions, and the implications for efficacy under field conditions are therefore not clear. In addition, few studies have examined the efficiency of foliar Mo fertilization in soybean-maize succession systems. The results of the present study show that foliar fertilization with Mo can modulate NR activity and processes involved in photosynthesis in soybean and maize, resulting in increased GY.

In general, foliar application of Mo improved the physiology and productivity of the studied crops. As expected, leaf N concentration was significantly increased in both crops, as Mo is directly involved in N metabolism and acts on important enzymes such as nitrogenase, xanthine dehydrogenase and NR, which are involved in N absorption, transport and conversion of NO₃⁻ to NO₂⁻, respectively (Bittner, 2014). In particular, the activity of NR, mainly its active form, increased, suggesting that the localized application of Mo to leaves effectively increased the expression and activation of NR, resulting in greater N assimilation by the crops. The absence of foliar Mo supplementation favored the accumulation of nitrate in leaves, indicating the low efficiency of the assimilation of nitrate under low concentrations of this micronutrient (Calonego et al., 2010). In leaves, the activation of NR is dependent on photosynthesis and is inactivated when light intensity is reduced or the leaf concentration of CO₂ is low (Kaiser and Spill, 1991; Provan and Lillo, 1999). Therefore, increased photosynthesis may also contribute to an increase in the concentration of active NR. On the other hand, the reduction and assimilation of N will benefit the whole photosynthetic apparatus (enzyme production) and increase photosynthesis.

Applying Mo also improved S content by increasing N content. N interacts with S, and thus a deficiency of one element reduces the absorption and assimilation of the other. The absorption and assimilation of S depend on a constant supply of the cysteine precursor, O-acetylserine, which in turn depends on an adequate supply of N (Abdin et al., 2003).

The increased Mo content in soybean and maize leaves indicated that this nutrient was efficiently absorbed by leaves. Absorbed Mo can be translocated to other organs via the phloem to reach, for example, the nodules of soybean plants within 5 days of application (Campo and Hungria, 2002; Steiner and Zoz, 2015).

The range of Mo sufficiency in the leaf tissue is 1–5 mg kg⁻¹ for soybean and 0.1–1 mg kg⁻¹ for maize (Walsh and Beaton, 1975; Novais et al., 2007). In all treatments, Mo was sufficient for both crops, but supplementation with Mo improved photosynthetic activity by increasing net photosynthesis (A) and stomatal conductance (gs) and reducing Ci. These improvements, even in well-nourished plants, support the use of stimulant foliar fertilization for plants with high yield potential to correct nutrient composition and improve plant metabolism (Rosolem, 2002). In summary, foliar fertilization with Mo may maximize physiological processes of the plant such as photosynthesis, ultimately leading to increased GY.

Typically, gs is used to measure the capacity of stomatal opening and closing in response to environmental stimuli, whereas Ci and A are used to measure the efficiency of carbon fixation by plants (Farquhar and Sharkey, 1982). Stomata are responsible for the exchange of carbon and water between the interior of the leaf and the atmosphere, processes that are fundamental to photosynthesis (Gago et al., 2016). Thus, the greater the gs, the greater the diffusion of CO₂ into the leaf, and this, combined with low levels of Ci, indicates that the mesophyll cells were able to assimilate CO₂ retained in the substomatal chamber through photosynthesis (Lawson and Blatt, 2014).

The increase in photosynthetic activity mediated by Mo supplementation may be due to two main effects: indirect effects of Mo on leaf N supply due to the participation of Mo in the synthesis of proteins such as Rubisco and direct effects due to the roles of Mo in chlorophyll biosynthesis and the stability of the photosynthetic apparatus (Yu et al., 2006; Carmo-Silva et al., 2015; Steiner and Zoz, 2015; Imran et al., 2019). Several studies have confirmed that the application of Mo improves chlorophyll content (Bambara and Ndakidemi, 2009; Wu et al., 2017; Li et al., 2018; Liu et al., 2020; Rana et al., 2020). In addition, Mo spraying has been reported to increase the number of chloroplasts and improve the stability of the cell wall and plasma membrane of thylakoids (Weng et al., 2009). Mo may also be involved in cytochrome b₆-f stability (Sun et al., 2014) and in improving chloroplast ultrastructure (Imran et al., 2019).

The improvement in photosynthetic activity was accompanied by increased WUE in maize, as leaf-level WUE reflects the ratio of net photosynthesis to transpiration (A/E). Since E was not affected by Mo application, carbon fixation increased under the same water use when Mo was applied (Figures 4I,J). During the time period of the study, significant dry spells occurred, especially in the second maize growing season (Figures 1B,D). The considerable increase in WUE in the presence of Mo compared with the control suggests that this element can improve tolerance to water deficit, ensuring an increase in photosynthetic and productive components even under water restriction. WUE is an important component of agricultural sustainability, especially in light of predictions of more frequent episodes of climatic anomalies such as droughts and heatwaves (Bhattacharya, 2019).

The increased protein content in soybean leaves may be the result of increased N and S content and photosynthetic activity. Moreover, increased N content is a strong indication that Mo application enhances N absorption and translocation. N is an essential structural element of proteins and accounts for 16%–23% of the content of plant proteins (Yeoh and Wee, 1994). In leaves, the most abundant protein is the enzyme Rubisco, which represents up to 50% of total soluble protein (Carmo-Silva et al., 2015). Increased protein content may therefore indicate an increase in Rubisco content. In support of this hypothesis, Maekawa and Kokubun (2005) observed a positive correlation between Rubisco content in soybean leaves and the applied dose of N. In addition, increased photosynthesis due to increased Rubisco levels has the potential to create a positive feedback effect by stimulating NR activation, which favors the assimilation of N and the onset of protein translation (Kaiser and Spill, 1991; Tcherkez et al., 2020).

Mo application did not alter PEPcase enzyme activity in maize leaves but did increase Rubisco enzyme activity in both crops. Mo fertilization has been reported to promote Rubisco stability under abiotic stress conditions (Sun et al., 2014). Together, the increased stability of Rubisco and higher protein content under Mo application could explain the improvement in the activity of this enzyme under drought stress. The increase in Rubisco enzyme activity led to an increase in total soluble sugar content, reflecting improved photosynthetic efficiency (Steiner and Zoz, 2015). Based on these results, it can be argued that Mo increases total soluble sugar by regulating photosynthesis. The high leaf soluble sugar content promoted by Mo fertilization may have been transported from the source to sinks via the phloem, resulting in increased NGP. Carbohydrate allocation differed between soybean and maize, as the number of pods per soybean plant increased, while maize did not change prolificacy. By contrast, the number of kernels per maize ear increased, whereas the number of kernels per soybean pod did not. Overall, grain yield increased by approximately 500 kg ha⁻¹ for both crops. This cascade of events culminated in a greater production of SDM which, associated with an increase in SNC, led to greater SNA. These results show how the localized application of micronutrients in small concentrations can change the entire metabolic mechanism of the plant, reflecting greater biological production.

Several studies of Mo fertilization have attributed increased GY to improvements in biological nitrogen fixation and NR activity (Campo et al., 1999; Calonego et al., 2010; Mercante et al., 2011; Steiner and Zoz, 2015; Crusciol et al., 2019). In the present study, supplementation with Mo improved GY not only by increasing the absorption and assimilation of N but also by altering photosynthetic properties, despite conditions of water scarcity in the two growing seasons, especially for maize. This response is a strong indication that foliar-applied Mo may be an important strategy not only to supply nutrients but also to stimulate carbon metabolism.

In maize, drought stress during flowering and emission of stigma styles can greatly reduce the number of grains and grain yield (Magalhães and Durães, 2006). Here, Mo fertilization increased the number of grains and maize yield, in addition to WUE, despite drought stress during the reproductive stage in both growing seasons. Although these findings represent a great advance in the understanding of the nutritional effects and stimulant action of Mo, many gaps remain, especially in relation to its anti-stress role. Mo influences the synthesis of abscisic acid (ABA), a hormone responsible for stomatal control under stress, and may be involved in the modulation of antioxidant enzymes (Watanabe et al., 2018). However, the effects of Mo on ABA have not been tested under field conditions and will be explored in further studies to confirm the role of Mo in coping with climate adversity.

In the present study, foliar Mo fertilization improved the physiology and productivity of soybean and maize crops by increasing NR activity, which in turn promoted increased leaf N content and protein synthesis. Foliar Mo fertilization also improved photosynthetic parameters, suggesting that foliar Mo fertilization is a viable strategy not only for promoting the nutritional status of the plant but also for stimulating carbon metabolism.

No adverse effects of Mo application were observed in the present study. Conversely, the positive effects of foliar fertilization with Mo were clear with the application of only 30 g Mo ha⁻¹. Both soybean and maize commercial crops are sprayed several times during their cycles for other nutrients and agrochemicals. Mo may be included in one of these spray campaigns with little or no additional cost but with significant potential benefits. Therefore, it is suggested that foliar Mo fertilization be included in soybean and maize nutrient management.

The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.

CC, JC, and HC designed the experiment. VR and SO obtained and processed the data. JP, JB, and TG analyzed the data. SO, LM, JC, JP, JB, and TG wrote the paper with contribution of all authors. All authors confirm being contributor of this work and has approved it for publication.

This study was financed by the National Council for Scientific and Technological Development (CNPq), providing the scholarship to the SO (grant number: 130646/2019-9).

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.