Ion-exclusion/cation-exchange chromatography using dual-ion-exchange groups for simultaneous determination of inorganic ionic nutrients in fertilizer solution samples for the management of hydroponic culture

https://doi.org/10.3390/agronomy11091847

“In this study, ion-exclusion/cation-exchange chromatography (IEC/CEC) using dual-ion-exchange groups (carboxy and sulfo groups) for the simultaneous determination of anions (SO₄²⁻, Cl⁻, NO₃⁻, and HPO₄²⁻) and cations (Na⁺, NH₄⁺, K⁺, Mg²⁺, and Ca²⁺) was developed. By using the combination of dual-ion-exchange groups, simultaneous separation of inorganic ions with HPO₄²⁻ was achieved that was impossible by the conventional IEC/CEC based on the single-ion-exchange group (carboxy group). This method was applied to the monitoring of inorganic ionic nutrients in fertilizer solution samples in hydroponic culture. As a result, a higher peak resolution of inorganic anions and cations with phosphate ion using IEC/CEC with dual-ion-exchange groups was achieved in the absence of matrix effects. In addition, the developed method helps to understand the behavior of ionic nutrients in fertilizer solution during hydroponic cultivation and is potentially useful for the individual fertilization of ionic nutrients.”

2.4. Hydroponic System

A closed hydroponic system (Green Farm UH-A01E1, Uing Co. Ltd., Osaka, Japan) was used as shown in Figure 1. In this study, Lactuca sativa was selected as the target crop, because of it is one of the major crops in greenhouse horticulture, including the closed hydroponic cultivation, in the world [16,17,18]. The fertilizer solution was prepared using 1.5 and 1.0 g/L of OAT House S1 and 2 fertilizer powders (OAT Agrio Co. Ltd., Tokyo, Japan) respectively, and diluting them with ultra-pure water. The fertilizer solution samples were collected on days 1, 4, 7, 9, 11, 14, 16, 21, 23, 25, and 28 from the hydroponic culture. After filtration through a polytetrafluoroethylene syringe filter with a 0.2 μm pore size (DISMIC^®-25HP, Advantec Toyo Kaisha, Ltd., Tokyo, Japan), the samples were temporarily refrigerated at 5 °C, diluted 5-fold, and immediately injected into the developed IEC/CEC system.

Figure 1. Closed hydroponic system used in this study. (A) Photo of the whole system, (B) storage of fertilizer solution with bubbling system and water volume meter, (C) seeding panel and support with seeds of Lactuca sativa, and (D) LED lighting system of the hydroponic system.

3.2. Eluent Optimization

The optimization of the tartaric acid concentration in the eluent with 2 mM 18-crown-6 was conducted to achieve a good separation of analyte ions using the IEC/CEC system. As shown in Supplementary Figure S3A, the retention times of the anions were slightly increased by enhancing the penetration effects, a side effect of the IEC, into the cation-exchange phase [19,20]. In contrast, the retention times of the cations dramatically decreased by increasing the tartaric acid concentration, which, in turn, reduced the cation-exchange effect. Hence, a higher concentration of tartaric acid in the eluent should be used to reduce the analytical time as much as possible. However, when the tartaric acid concentration was increased from 2 to 8 mM, the peak resolution (Rs) between the Na⁺–NH₄⁺ (1.16–0.67) and the NH₄⁺–K⁺ (1.82–0.97) was inadequate, as shown in Supplementary Figure S4A. Hence, 5 mM of tartaric acid was used as the optimal concentration because it afforded the best resolution of the analyte ions under the maximum possible reduced analytical time.

Next, varying concentrations of 18-crown-6 were added to 5 mM of the tartaric acid eluent to improve the resolution between NH₄⁺ and K⁺ [21]. The separation is based on the stability constant of the complexation of alkali metal ions with 18-crown-6 (log K_Na+ = 0.80, log K_NH4+ = 1.23, and log K_K+ = 2.03) [22]. When included in the eluent, 18-crown-6 absorbs onto the cation-exchange resin in the column and increases the retention time of the associated monovalent cations [23]. The retention time of K⁺ increased depending on the 18-crown-6 concentration, as shown in Supplementary Figure S3B. The peak resolution (Rs) between NH₄⁺ and K⁺ also improved when the concentration of 18-crown-6 increased in the following order: 1 mM (Rs = 0.60) < 2 mM (Rs = 1.34) < 3 mM (Rs = 2.02) < 4 mM (Rs = 2.33) < 5 mM (Rs = 2.65) < 6 mM (Rs = 3.10), as shown in Supplementary Figure S4B. The 3 mM concentration of 18-crown-6 was selected as the optimal concentration because the obtained Rs value was higher than the 1.5 required to ensure total separation.

Consequently, the IEC/CEC using 5 mM tartaric acid and 3 mM 18-crown-6 as the eluent (pH 2.98) successfully achieved simultaneous separation of the analyte ions with phosphate ion, as shown in Figure 2A. In addition, the optimized system could separate the inorganic ionic components in 5-fold diluted fertilizer solution collected on day 9 of the hydroponic cultivation, as shown in Figure 2B.

Figure 2. Optimal chromatograms for the separation of (A) standard sample, and (B) 5-fold diluted fertilizer solution sample collected on day 9. Peaks: (1) SO₄²⁻, (2) Cl⁻, (3) NO₃⁻, (4) H₂PO₄⁻, (5) Na⁺, (6) K⁺, (7) NH₄⁺, (8) Mg²⁺, and (9) Ca²⁺. Conditions: Column: Two Tosoh TSKgel Super-IC-A/C (6.00 mm I.D. × 150 mm) and a Shodex SH-1011 (8.00 mm I.D. × 300 mm) connected in tandem; column temperature: 55 °C; injection volume: 20 µL; eluent concentration: 5 mM tartaric acid and 3 mM 18-crown-6; eluent flow rate: 1.0 mL/min. Injected samples: (A) mixture of 1.0 mM MgSO₄, NaCl, NH₄NO₃, KH₂PO₄, and CaCl₂, and (B) 5-fold diluted fertilizer solution sample collected on day 9.