4.1 The effects of foliar arginine on plant growth, fruit yield and quality
In this study, tomato plants treated with arginine showed increased plant growth, fruit yield, and fruit quality compared with the control plants. Foliar application significantly promoted plant biomass and stem diameter. Notably, arginine treatment resulted in increased tomato fruit weight, size, and yield per plant. Additionally, exogenous arginine increased the concentrations of lycopene, vitamin C, and soluble protein; the contents of soluble sugar, titratable acid, and soluble solids; and the sugar-acid ratio.
We showed that arginine induced increased growth of the aboveground plant parts and increased root development (Fig. 1). A previous study indicated a similar positive effect of arginine on plant growth in which the growth of bean seedlings was improved by the application of arginine (Zeid, 2009). Naser and Azeez (2019) found that the stem diameters and leaf areas of apricot trees were significantly increased following spraying with arginine. Our results showed that tomato fruit yield, weight, and size were significantly increased following spraying with arginine (Fig. 4). These results match those observed in earlier studies showing improved yields following arginine application. Abdul-Qados (2009) concluded that spraying plants with arginine effectively increased wheat growth and all yield components. Mohseni et al. (2017) demonstrated that for strawberry, the yield, the mean weights of primary and secondary fruits, and the number of achenes were increased following foliar application of arginine.
Our study also found that the exogenous application of arginine significantly enhanced tomato quality. Foliar application of arginine resulted in increased concentrations of lycopene and vitamin C in tomato fruits (Fig. 5a, b). Lycopene and vitamin C are vital for the nutritional quality of tomato fruits (Frusciante et al., 2010). In recent years, lycopene has been increasingly used in the food industry and in medicines, and has shown benefits for human health (Grabowska et al., 2019). One study showed that the lycopene concentration in tomato fruits declined with nitrogen fertilizer supplementation (Dumas et al., 2003). However, our data contradict this finding, and this difference is difficult to explain due to a lack of evidence in the literature. Further investigation is needed to clarify the reasons behind the differing findings. Vitamin C deficiency is a crucial issue affecting people’s health and results in scurvy (Brickley et al., 2020). A previous study indicated that nitrogen application increased the vitamin C concentration in leafy parsley (Koota, 2011). Other studies have also reported increases in vitamin C concentration in various crops following the application of amino acids (Noroozlo et al., 2019; Mohammadipour and Souri, 2019; Souri et al., 2017; Souri and Hatamian, 2019; Shooshtari et al., 2020). Thus, the enhanced vitamin C concentration in tomato fruits could be due to the organic nitrogen supplied by the foliar arginine treatment.
Soluble solids, soluble sugar, and titratable acids are important components of flavor in tomato fruit (Atanassova et al., 2003). Several studies showed that soluble solids are related to nitrogen supplementation in various plants (He et al., 2003; Pierre et al., 2008). Our results showed that soluble sugar, titratable acids, and the sugar-acid ratio in tomato fruits were promoted by exogenous arginine (Fig. 5c, d, e). Our findings agree with Mohseni et al. (2017), who demonstrated significantly increased induction of sugars and titratable acids in strawberry following arginine application. Recent evidence has shown that arginine plays a vital role in organic acid metabolism in plants (Hildebrandt et al., 2015). Another crucial, and novel finding was that the sugar-acid ratio was higher than the control, indicating that the tomato became sweeter following external arginine application.
4.2 Mechanisms by which arginine improves tomato plant and fruit growth
Our study found that foliar application of arginine resulted in a significant improvement in the rate of photosynthesis in tomato leaves, and this was also seen in previous studies (Stephan et al., 2000). Yagi and Al-Abdulkareem (2006) demonstrated that arginine showed positive effects on chlorophyll synthesis and improve photosynthesis in Eruca sativa Mill. Marschner (2011) illustrated that carbohydrates accumulated via photosynthesis formed the basic framework of plants. Increased photosynthesis following the external application of amino acids has been identified as a major factor that promotes plant growth (Yagi and Al-Abdulkareem, 2006; Mohammadipour and Souri, 2019; Noroozlo et al., 2019). Petridis et al. (2018) reported that increased tomato fruit yield was mainly associated with improved carbohydrate content due to increased photosynthesis. Meanwhile, the increased accumulation of soluble solids and soluble sugar in arginine-treated tomato fruits was consistent with the increase in carbohydrates. Arginine induced photosynthesis, therefore, might be responsible for the increased tomato fruit yield and quality. Nitrogen is vital for photosynthesis as it participates in chlorophyll synthesis, chloroplast structure stabilization, and the activation of relevant enzymes (Marschner, 2011). It is likely that the increased photosynthetic rate we observed in tomato leaves could be related to the increased nitrogen (Fig. 2). In summary, exogenous arginine increased the nitrogen concentration leading to improved photosynthesis in tomato leaves, resulting in a significant increase in growth of the aboveground plant parts and the roots, as well as fruit yield and quality.
In plants, arginine is regarded as an organic nitrogen storage sink and nitrogen transformation medium due to its high nitrogen-to-carbon ratio (Winter et al., 2015). It has been reported that amino acids can be absorbed and utilized directly as organic nitrogen sources by plants (Nasholm et al., 2009; Ganeteg et al., 2017). Jonas and Torgny (2001) demonstrated that pine trees assimilate arginine and showed a similar response to treatment with ammonium and nitrate. In recent years, improved nitrogen utilization in plants following arginine application has been well documented (Abdul-Qados et al., 2009; Ghasemi et al., 2012). We found a similar effect on nitrogen accumulation in tomato, which is likely due to arginine uptake. In our experiment, 50 ml of 1 mmol/L arginine was applied 12 times per tomato plant, resulting in a total application of 104.52 mg of arginine during the experimental period. We calculated that the tomato plants absorbed a maximum of 33.6 mg of nitrogen with arginine treatment. However, the nitrogen content in the aboveground parts of the tomato plants was increased by 0.3 g compared with the control plants (Figure S1). Obviously, the nitrogen supplied by the direct uptake of arginine was insufficient for the nitrogen increment observed in the aboveground parts. Therefore, other factors may be involved in the improved nitrogen accumulation in the arginine-treated tomato plants.
In plants, nitrogen uptake is mainly regulated by the gene families encoding nitrite transporter (NRT) and ammonium transporter (AMT) proteins. Moreover, LeNRT1.1 and LeAMT1.1 are reported to be the primary regulatory genes for the uptake of nitrate and ammonium in tomatoes (Lauter et al., 1996; Filiz and Akbudak, 2020). Nitrogen metabolism in plants is regulated primarily by nitrate reductase, glutamine synthetase, and glutamate synthase, encoded by the LeNR, LeGS1, and LeGOGAT genes, respectively (Becker et al., 1993; Yao et al., 2008). Compared with the control, the expression of LeNRT1.1 was upregulated markedly by arginine treatment in our study (Fig. 2c). However, we found no differences in the expression of LeAMT1.1, LeNR, LeGS1, or LeGOGAT (Figure S2). Hence, we conclude that arginine has a positive effect on nitrate uptake but not on ammonium uptake and nitrogen transport in tomato plants. Our findings indicate a potential second mechanism underlying the increased nitrogen concentration in the tomato plant aboveground parts. However, further research is needed to fully determine the relationship between external arginine application and the nitrate absorption pathway in tomato.
Studies have shown that arginine is essential for root growth and elongation, and other amino acids cannot replace arginine (Xia et al., 2014a, b). White et al. (2015) demonstrated that increased root development in grass enhanced the capacity to absorb nitrogen. Additionally, the promotion of root activity had a positive effect on nitrogen uptake in tomato plants under root-zone heating conditions (Kawasaki et al., 2014). In our study, the significant increase in root activity and plant growth following arginine treatment helped tomato plants assimilate nitrogen from the soil (Fig. 1c, d). Hence, arginine application leading to increased root growth might underlie the enhanced accumulation of nitrogen in the tomato aboveground plant parts and in the fruit.
In summary, the increase in nitrogen seen in the aboveground parts and fruit of the tomato plant is related to three possible arginine-induced mechanisms (Fig. 5). These mechanisms involve arginine uptake as organic nitrogen, increased LeNRT1.1 expression, and increased root development and activity. It is possible that a small amount of arginine applied as a foliar spray could result in considerable nitrogen accumulation in the tomato plant and fruit. This increased nitrogen concentration significantly promotes photosynthesis in the tomato leaves, which stimulates plant growth. Moreover, tomato fruit yield, flavor, and nutritional quality were directly improved by the exogenous application of arginine. In addition, arginine is a main component of proteins found in living organisms and does not harm the environment, making it an excellent candidate for agricultural application.