Hydrogen-Rich Water Alleviates Aluminum-Induced Inhibition Of Root Elongation In Alfalfa (Medicago Sativa L.) VIA Decreasing Nitric Oxide Producing

Alfalfa (Medicago sativa L.) is a perennial forage and it belongs to the legume plant. the crude protein content of its early flowering stage is 16% to 20%, so it’s rich in nutritional value. The distribution of alfalfa in the world is very wide and it plays extremely important role in agriculture and animal husbandry. Excessive aluminum (Al) in acidic soil causes toxic damages to plants, of which the most obvious is the inhibition of primary root elongation. Previous studies showed that nitric oxide (NO) is involved in plant responses to metal stress. In recent years, the studies on animals found that hydrogen (H2) plays selective anti-oxidative, anti-inflammatory and anti-stress roles. However, there is no evidence proving whether H2 is involved in plant aluminum toxic response. Therefore, the experiment was carried out to explore how hydrogen-rich water (HRW) to ease aluminum stress on the inhibition of alfalfa primary root elongation. The results showed that:1. HRW pretreatment could alleviate Al-induced inhibition of primary root elongation in alfalfa seedlings. This study found that Al3+ significantly inhibited the elongation of primary root of alfalfa seedlings in acidic condition (pH 4.5), and it showed a concentration-dependent effect within a certain range. By further experiment, we chose the AlCl3 concentration (150 μM) of 50% inhibitory effect. When seedlings were exogenously pretreated with different saturations of HRW, the inhibition was differentially relieved, especially 50% saturation of HRW works best. Besides, the addition of 50% HRW also alleviated the appearance of Al toxicity symptoms, including the improvement of seedling growth and accumulated less amount of Al. However, in case of co-treatment, the relief was not obvious;2. Nitric oxide (NO) was involved in the inhibition of primary root elongation caused by Al stress in alfalfa seedlings. The pretreatment of NO scavenger PTIO and cPTIO could significantly alleviate Al-induced inhibition of alfalfa root elongation, the effect is similar with HRW. However, the effect of NO releasing compounds (SNP) is similar with Al, which could significantly inhibit root elongation.3. HRW alleviated Al-induced inhibition of primary root elongation by decreasing endogenous NO content. The results of the different combination treatments with PTIO, SNP and HRW have shown that the reduction of the endogenous NO can effectively alleviate Al stress. The results of laser scanning confocal microscopy (LSCM) and electron paramagnetic resonance(EPR) verified that the exogenous HRW pretreatment could significantly reduce Al-induced NO accumulation in roots of alfalfa seedlings, and its effect is similar to that of NO scavengers, but contrary to NO releasing compound. The Al-induced inhibition of root elongation was markedly recovered by tungstate, an inhibitor of the NO synthetic enzyme nitrate reductase (NR), and the effect was similar with HRW; it suggested that NR activity might be implicated in this response.Thus, these above studies have shown that HRW alleviated Al-induced inhibition of alfalfa primary root elongation by decreasing NO production.