Hijaz Faraj, Nehela Yasser, Batuman Ozgur, Killiny Nabil
Citrus Research and Education Center, Department of Plant Pathology, IFAS, University of Florida, 700 Experiment Station Road, Lake Alfred, FL 33850, USA.
Department of Agricultural Botany, Faculty of Agriculture, Tanta University, Tanta 31512, Egypt.
Antibiotics (Basel). 2021 May 12;10(5):566. doi: 10.3390/antibiotics10050566.
Oxytetracycline (OTC) and streptomycin have been used for the control of several plant diseases and were recently permitted for the control of citrus greening disease, Huanglongbing. Consequently, sensitive and reliable methods are highly needed for the detection of OTC in citrus tissues. Herein, we studied the replacement of cetyltrimethylammonium chloride (CTAC) by citrate (Cit) as a sensitizing agent for the analysis of OTC in citrus tissues using the recently established europium (Eu) method. In addition, we determined the optimal conditions for the formation of the Eu-OTC-Cit ternary complex in tris buffer. Our results showed that the plant matrix significantly decreased the fluorescence intensity of the Eu-OTC-Cit complex even after the replacement of CTAC. Our investigations showed that phenols such as gallic acid degrade slowly at high pH and their degradation was enhanced in the presence of the (Eu) cation. To reduce the plant matrix interference, the sample extract was cleaned using solid-phase extraction (SPE). The OTC recoveries from spiked healthy and Liberibacter asiaticus (Las)-infected trees were 91.4 ± 7.8% and 82.4 ± 3.9%, respectively. We also used the citrate method to determine the level of OTC in trunk-injected trees. The level of OTC as measured using the Eu-OTC-Cit complex (117.5 ± 20.3 µg g fresh weight "FWT") was similar to that measured using Eu-OTC-CTAC complex (97.5 ± 14 µg g FWT). In addition, we were able to visualize the OTC in citrus leaf extract, under ultraviolet light (400 nm), after it was cleaned with the SPE. Our study showed that the citrate can be successfully used to replace the harmful CTAC surfactant, which could also react with phenols.
土霉素（OTC）和链霉素已被用于防治多种植物病害，最近被允许用于防治柑橘黄龙病。因此，迫切需要灵敏可靠的方法来检测柑橘组织中的OTC。在此，我们研究了用柠檬酸盐（Cit）替代十六烷基三甲基氯化铵（CTAC）作为增敏剂，采用最近建立的铕（Eu）法分析柑橘组织中的OTC。此外，我们还确定了在三羟甲基氨基甲烷缓冲液中形成Eu-OTC-Cit三元络合物的最佳条件。我们的结果表明，即使在替代CTAC后，植物基质仍显著降低了Eu-OTC-Cit络合物的荧光强度。我们的研究表明，诸如没食子酸之类的酚类物质在高pH值下降解缓慢，并且在（Eu）阳离子存在下其降解会增强。为了减少植物基质干扰，使用固相萃取（SPE）对样品提取物进行净化。加标健康树和感染亚洲韧皮杆菌（Las）的树中OTC的回收率分别为91.4±7.8%和82.4±3.9%。我们还使用柠檬酸盐法测定了树干注射树木中OTC的含量。使用Eu-OTC-Cit络合物测得的OTC含量（117.5±20.3 µg g鲜重“FWT”）与使用Eu-OTC-CTAC络合物测得的含量（97.5±14 µg g FWT）相似。此外，在用SPE净化后，我们能够在紫外光（400 nm）下观察到柑橘叶提取物中的OTC。我们的研究表明，柠檬酸盐可以成功地用于替代有害的CTAC表面活性剂，CTAC也会与酚类物质发生反应。
