Ion [35]. The MDA content material at 532 nm was calculated by subtracting the absorbance at 600 nm. 2.5. Leaf Photosynthesis, Chlorophyll fluorescence Parameters, and Chlorophyll Content material The net photosynthetic price (Pn), stomatal conductance (Gs), transpiration rate (Tr), and intercellular CO2 concentration (Ci) from the leaves were measured by the portable photosynthetic program (li-6400, Li-COR, Lincoln, NE, USA). Leaf photosynthetic parameters were determined at 10 a.m. following the plants had been treated with distinctive Fexinidazole Epigenetic Reader Domain concentrations of NaCl and treated with unique concentrations of calcium chloride for 1 week. The mature leaves had been dark-adapted for 20 min without the need of isolation, and also the fluorescence kinetic parameters at space temperature had been measured making use of a portable modulation chlorophyll fluorescence instrument (PAM-2500 Walz, Effeltrich, Germany). For the chlorophyll content, 0.03 g of fresh leaves were extracted in a 10 mL pigment extraction resolution containing absolute ethanol and acetone (1:two, v/v) at 25 C for 12 h inside the dark. The absorbance on the supernatant at 470, 645, and 663 nm was then measured working with an ultraviolet spectrophotometer. Chlorophyll a, chlorophyll b, carotenoids, and total chlorophyll content material were calculated according to [36]. 2.6. Determination of K+ , Na+ , and Ca2+ To establish the K+ , Na+ , and Ca2+ ion concentrations, we meticulously washed fresh root, stem, and leaf samples with deionized water, placed them in an oven at 105 C for 20 min, after which kept the temperature continuous at 80 C until the samples were entirely dried. The dried plant samples were then grounded in a 5 mL centrifuge tubes utilizing a high-throughput plant tissue ball milling instrument (Scientz-192, Xinzhi Biotechnology Co., Ltd., Ningbo, China). A total of 0.3 g of every sample powder was weighed, and five mL of nitric acid and 1 mL of perchloric acid have been added for wet digestion. The K+ , Na+ , and Ca2+ contents of plant tissue extracts and regular samples (National Institute of Metrology, Beijing, China) had been determined by inductively coupled plasma optical emission spectrometer (ICP-OES; PerkinElmer, Optima 8300, Waltham, MA, USA). The concentration of K+ , Na+ , and Ca2+ is defined as K+ , Na+ , and Ca2+ content material (mg) per unit tissue (g) [37]. 2.7. Extraction and LC S Evaluation of Phenolic Compounds 2.7.1. Chemical substances and Reagents UPLC-grade acetonitrile and methanol were bought from Fisher Scientific (Pittsburgh, PA, USA). All other reagents were of analytical purity. Ultrapure water was prepared by a Milli-Q system (Millipore, Bedford, MA, USA) water 12-Hydroxydodecanoic acid MedChemExpress purification method. The reference compounds required for the experiment had been all purchased from ChromaDex Inc. (Santa Ana, CA, USA), such as p-hydroxycinnamic acid, p-hydroxybenzoic acid, two,5-dihydroxybenzoic acid, genistein, abscisic acid, petunidin, naringenin, hesperidin, quercetin-3-O-rhamnoside, chlorogenic acid, ferulic acid, myricetin, luteolin, catechin, cinnamic acid, p-coumaric acid, hesperetin, quercetin, caffeic acid, L-phenylalanine, naringin, kaempferol, liquiritigenin, isoliquiritigenin, and vanillic acid. The purities of those requirements were greater than 98 .Agriculture 2021, 11,5 of2.7.two. Preparation of Test Sample Resolution Gleditsia sinensis plant tissues (root, stem, and leaf) treated with different treatments (CK, S1, S2, S1 + C1, S1 + C2, S1 + C3) had been grounded after which ultrasonically extracted (100 kHz, 40) for 45 min by adding 10 mL of 70 methanol. Immediately after filtration, the.