Amberlite irp 64 cation exchange resin

The MALDI-TOF MS spectra were recorded on Bruker Reflex III (Germany). The experiment was carried out with reference to the method described by Xiang et al. (2006) [32] . 2,5-Dihydroxybenzoic acid (DHB, 10 mg/ml 30% acetone solution) was used as matrix. Amberlite IRP-64 cation-exchange resin (Sigma-Aldrich, USA), equilibrated in deionized water, was used to deionize the analyte, matrix, and cesium chloride solution. The sample solutions (10 mg/ml 30% acetone solution) were mixed with the cesium chloride (1.52 mg/ml) solution at a volumetric ratio of 1∶1 to promote the formation of a single type of ion adduct ([M+Cs]⁺). Next, the mixture was mixed with the matrix solution (1∶3, v/v). Then, the mixture (1 μl) was spotted to the steel target.

Water used in this experiment was purified on a Millipore Milli-Q apparatus. HPLC grade dichloromethane, acetonitrile (CH₃CN), methanol, trifluoroacetic acid (TFA), acetic acid, and all analytical grade solvents (acetone, methanol, n-Butanol etc.) were obtained from Sinopharm (Sinopharm, Shanghai, China). Sephadex LH-20, Folin-Ciocalteu reagents, acetone-d₆, deuteroxide (D₂O), Amberlite IRP-64 cation-exchange resin, cesium chloride (CsCl), 2,5-dihydroxybenzoic acid (DHB), benzylmercaptan, 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,4,6-tripyridyl-S-triazine (TPTZ), ascorbic acid (AA), and all HPLC standards were purchased from Sigma (St. Louis, MO, USA). The mature leaves (the third pair with fully expanded and dark green) of a mangrove plant C. tagal (Rhizophoraceae Ceriops), were collected from a mangrove forest in the Dongzhai harbor (19°56′N, 110°34′E), Hainan, China. The leaves were immediately freeze-dried, ground, and stored at −20°C prior to analysis.

MALDI-TOF-MS was performed according to the method described by Zhang et al.16 (link). The MALDI-TOF-MS spectra were recorded on a Bruker Reflex III instrument (Germany). The irradiation source was a pulsed nitrogen laser with a wavelength of 337 nm, and the duration of the laser pulse was 3 ns. In the positive reflection mode, an accelerating voltage of 20.0 kV and a reflectron voltage of 23.0 kV were used. Spectra of PAs were obtained from a sum of 100–150 shots and calibrated using angiotensin II (1,046.5 MW), bombesin (1,619.8 MW), ACTHclip18–39 (2,465.2 MW), and somatostatin 28 (3,147.47 MW) as external standards. 2,5- dihydroxybenzoic acid (DHB, 10 mg/mL aqueous solution) was used as the matrix. The sample solutions (7.5 mg/mL aqueous) were mixed with the matrix solution at a volumetric ratio of 1:3. The mixture (1 μL) was applied to the steel target. Amberlite IRP-64 cation-exchange resin (Sigma-Aldrich), equilibrated in deionized water, was used to deionize the analyte/matrix solution three times. Cesium chloride (1 mg/mL) was mixed with the analyte/matrix solution at 1:3 volumetric ratio to promote the formation of a single type of ion adduct ([M + Cs]⁺)68 69 .