Iodine Biofortification of Dandelion Plants (Taraxacum officinale F.H. Wiggers Coll.) with the Use of Inorganic and Organic Iodine Compounds

Iodine is a crucial microelement necessary for the proper functioning of human and animal organisms. Plant biofortification has been proposed as a method of improving the iodine status of the population. Recent studies in that field have revealed that iodine may also act as a beneficial element for higher plants. The aim of the work was to evaluate the efficiency of the uptake and accumulation of iodine in the plants of dandelion grown in a pot experiment. During cultivation, iodine was applied through fertigation in inorganic (KI, KIO3) and organic forms (5-iodosalicylic acid, 5-ISA; 3,5-diiodosalicylic acid, 3,5-diISA) at two concentrations (10 and 50 µM). The contents of total iodine and iodosalicylic acids, as well the plant biomass and antioxidant capacity of dandelion leaves and roots, were analyzed. The uptake of inorganic and organic forms by dandelion plants was confirmed with no negative effect on plant growth. The highest efficiency of improving iodine content in dandelion leaves and roots was noted for 50 µM KI. The applicability of iodosalicylates, especially 5-ISA, for plant biofortification purposes was confirmed, particularly as the increase in the iodine content after the application of 5-ISA was higher as compared to that with commonly used KIO3. The chemical analyses have revealed that iodosalicylates are endogenous compounds of dandelion plants.


Introduction
Iodine is a crucial element for the proper functioning of human and animal organisms. Its necessity results from its presence in thyroid hormones: triiodothyronine and thyroxine. The recommended daily intake of iodine has been established on the following levels: 90 µg I for 0-6 yo children, 120 µg I for 6-12 yo children, 150 µg for adults and 250 µg I for pregnant and lactating women [1]. The insufficient supply of iodine may lead to, among others, malfunction of the thyroid gland, increased risk of pregnancy loss, growth retardation and neurological deficits grouped as IDDs-iodine deficiency disorders [1]. The main sources of iodine in a daily diet include: table salt, dairy products, bread, meat and fish [2]. In the coastal areas, iodine deficiency is less likely to occur due to the substantial consumption of marine fish that are naturally rich in that element. In the terrestrial regions, however, the problem of the insufficient supply of iodine needs to be monitored pro-health, including antioxidant, anti-inflammatory, hypoglycemic and hepatoprotective properties [30,31]. Despite its popularity, no studies on the mineral biofortification of dandelion have been conducted so far.
The primary aim of the work was to evaluate the possibility of improving iodine content in dandelion plants through the root application of different doses of inorganic (KI and KIO 3 ) and organic (5-iodosalicylic acid, 5-ISA; 3,5-diiodosalicylic acid, 3,5-diISA) iodine forms. It was also aimed at describing the effect of iodosalicylates on plant growth and the content of selected antioxidative compounds that may act as stress indicators in plants.

Growth of Dandelion Plants
The application of tested inorganic and organic iodine compounds had no significant effect on the growth of dandelion plants, as measured by the weights of leaves and roots of a single plant ( Table 1). The dry matter content in leaves and roots was however affected. As compared to the control, an increase in dry matter content in leaves was noted for the combination of KI 50 µM and 3,5-diISA. The lowest values of that parameter were found in leaves of plants grown in the presence of KIO 3 at both applied doses (10 and 50 µM). As compared to the control combination, dry matter content in roots decreased significantly in plants with combinations of 50 µM doses of KI, 5-ISA and 3,5-diISA. Data (mean ± SE) in columns followed by the same letter are not significantly different at p < 0.05 (n = 4).

Iodine Content in Dandelion Roots and Leaves
Plant cultivation in the substrate enriched with tested inorganic and organic iodine compounds increased the total content of iodine in leaves and roots of dandelion plants ( Table 2). The highest total accumulation of that element in dandelion plants was found for the application of 50 µM KI: 133.5 mg I kg −1 and 19.1 mg I kg −1 in leaves and roots, respectively. A substantial level of iodine uptake was also noted after the application of a 50 µM I dose of 5-ISA (99.2 and 17.1 mg I kg −1 in leaves and roots, respectively). Between the two inorganic sources of iodine, namely KI and KIO 3 , a higher effect of iodine enrichment in both roots and leaves was noted for KI irrespective of the applied dose. In the case of organic sources of iodine, the significant differentiation between 5-ISA and 3,5-diISA, in terms of increasing the total I content in leaves and roots, was observed only for the higher dose (50 µM). The application of 50 µM 5-ISA led to a greater accumulation of total iodine in both plant parts (99.2 and 17.1 mg I kg −1 in leaves and roots, respectively) as compared to that with the same concentration of 3,5-diISA (38.5 and 9.1 mg I kg −1 in leaves and roots, respectively). The application of 50 µM 5-ISA, an organic source of iodine, allowed to obtain a higher level of iodine in dandelion plants (both leaves and roots), compared to the inorganic compound KIO 3 applied at the same dose. Table 2. The contents of total iodine, 5-iodosalicylic acid (5-ISA) and 3,5-diiodosalicylic acid (3, in dandelion leaves and roots.

Combination
Leaves Roots The contents of 5-ISA and 3,5-diISA were analyzed in the leaves and roots of dandelion plants in all tested combinations (Table 2). There was little differentiation in the content of 5-ISA in dandelion leaves, and a significant increase in its level was noted only after the application of 50 µM 5-ISA. On the other hand, the content of 5-ISA in roots was not different from the control value in the case of both doses of KIO 3 and 10 µM KI. The highest level of that compound (6.3 ng g −1 ) was noted in the roots of plants grown in the substrate with 50 µM I 5-ISA. The applied dose of 3,5-diISA did not affect the level of 5-ISA in dandelion roots. Additionally, increasing the KI dose from 10 to 50 µM I led to a significantly higher content of 5-ISA in roots (0.1 and 0.8 ng g −1 , respectively). A significant increase in the leaf level of 3,5-diISA was noted only for the combinations with 3,5-diISA application, with the highest level noted for the 50 µM concentration: 8.88 ng 3,5-diISA g −1 .
In the case of roots, an increase in the 3,5-diISA level was noted for both doses of 3,5-diISA, as well as the 50 µM dose of 5-ISA. In general, the content of both iodosalicylates was higher in roots than in leaves of dandelion plants. Analyses of control plants showed that iodosalicylates are endogenous compounds of dandelion plants.

The Content of Phenolic Compounds and Antioxidant Capacity of Dandelion Plants
The application of inorganic and organic sources of iodine modified the level of phenolic compounds in dandelion leaves and roots (Tables 3 and 4). The highest total content of phenolics was noted in the leaves of plants grown in the presence of 50 µM KIO 3 (50.2 mg CAE g −1 ), while the lowest was in the leaves of plants from the combinations: 10 µM KI (37.2 mg CAE g −1 ) and 50 µM 5-ISA (36.5 mg CAE g −1 ). In all remaining combinations, the leaf accumulation of phenolic compounds did not differ significantly from the value noted for the control plants. The root content of total phenolics was the highest for the combinations with KI application (10.2 and 11.7 mg CAE g −1 , for 10 and 50 µM I, respectively). Increased root levels of these compounds, as compared to the control, were also noted for 50 µM doses of 5-ISA (9.0 mg CAE g −1 ) and 3,5-diISA (9.7 mg CAE g −1 ).
For most tested treatments, the introduction of iodine compounds into the substrate decreased the level of salicylic acid in plants, as compared to the control (Tables 3 and 4). Only in the case of 50 µM KI was an increase in the SA content in leaves observed (69.0 ng g −1 , Table 3). The increase in the applied iodine dose had no effect on the SA level in dandelion leaves, with the exception of higher SA content after the application of 50 µM KI, as compared to that with 10 µM KI. In the case of roots, the iodine dose variously affected SA accumulation depending on the tested compound (Table 4). For 50 µM KI and 3,5-diISA, there was a noted decrease, while for KIO 3 , there was an increase in the SA level as compared to respective combinations with lower dose of iodine. The applied dose of 5-ISA had no effect on SA accumulation in dandelion roots.  There was significant differentiation between the tested combinations with respect to the accumulation of jasmonic acid (JA) in dandelion leaves and roots (Tables 3 and 4). The introduction of inorganic and organic iodine sources into the substrate increased the content of JA in dandelion leaves, as compared to that in the control plants ( Table 3). The highest level of that compound was noted for treatments with both doses of 3,5-diISA (182.4 and 175.2 ng g −1 ), as well as the 50 µM dose of 5-ISA (164.6 ng g −1 ). The applied iodine dose had no significant effect on the leaf level of JA from KIO 3 , KI and 3,5-diISA treatments. Only in the case of 5-ISA application did an increase in the iodine dose lead to an increase in the JA level in dandelion leaves. The content of JA in dandelion roots decreased significantly, as compared to the control, after the application of KI, 5-ISA and 3,5-diISA (Table 4). Only the introduction of KIO 3 into the substrate led to a significantly increased level of JA in roots (432.0 and 439.3 ng g −1 ). Increasing the dose of iodosalicylates (5-ISA and 3,5-diISA) from 10 to 50 µM decreased the accumulation of JA in roots; such an effect was not noted for KI.
There was little differentiation observed in the antioxidant capacity of dandelion leaf extracts, as measured using the CUPRAC method ( Table 5). The highest CUPRAC values were noted in the leaves of plants fertigated with 3,5-diISA in both doses (214.6 and 215.1 µmol TE g −1 ). However, when individual combinations were taken into account, only in the case of 50 µM KI and 3,5-diISA applications did the obtained values differ significantly from the control. FRAP values of dandelion leaves increased significantly and similarly with almost all tested combinations, as compared to the control values, with the exception of 10 µM KIO 3 and 50 µM KI (Table 5). Additionally, the FRAP value for leaves with the 10 µM KIO 3 combination was significantly lower than those obtained when 5-ISA and 3,5-diISA were applied at both doses, as well as KI applied at a 10 µM KI dose. Little effect of iodine compounds was found with respect to the radical scavenging capacity of leaf extracts, as measured with the use of the DPPH method (Table 5). A significant increase in that parameter, as compared to the control, was noted only in plants fertigated with 50 µM KIO 3 (72.8 µmol TE g −1 ). The antioxidant capacity of dandelion roots, as measured using CUPRAC, FRAP and DPPH methods, increased significantly, as compared to the control, in combination with KI application at both doses ( Table 6). The lowest levels of FRAP values were noted in plants fertigated with 10 µM 3,5-diISA. An increase in the applied dose from 10 to 50 µM I led to a significant increase in the antioxidant potential, as measured by all used methods, in the roots of plants fertigated with 3,5-diISA. An increase in the KIO 3 dose contributed to significantly higher levels of CUPRAC and FRAP values of dandelion roots, while in the case of KI, a significant increase in the FRAP value was accompanied by the increased I dose. The application of a higher iodine dose of 5-ISA decreased the CUPRAC value of dandelion roots with no effect on the antioxidant potential, as measured via FRAP and DPPH methods.

Discussion
For years, iodine was not considered an important mineral element for plants. Only limited information was available considering the potential beneficial effect of that element on crop plant growth, while practical applications were limited to being a defoliating and herbicide agent [32]. In the last decade, however, an interest in that element has increased, in large part owing to the possibility of improving the human intake of iodine through the consumption of the so-called biofortified food. The on-going research on the potential of increasing the iodine content in edible plants has substantially improved the recognition of the role of iodine in higher plants. It has been discovered that relatively low doses of iodine not only can be neutral (non-toxic) for plants but also may increase their growth (for a review see [10,12]). The results obtained in the present study showed no significant effect of inorganic and organic sources of iodine, applied at two different but still low doses (10 µM and 50 µM I), with respect to the growth of dandelion plants in pot cultivation. This stays in general agreement with other studies reporting the effect of similar iodine doses during plant cultivation. No toxic effects on lettuce growth were noted when iodine (KI or KIO 3 ) was applied in the hydroponics nutrient solution in the 0-80 µM I concentration range [33]. The studies by Kiferle et al. [28] have revealed that KI and KIO 3 applied at a 100 µM I dose had no effect on the dry weight of basil plants grown in the soil. However, there is limited information on safe doses of iodine applied as organic compounds. Grzanka et al. [34] noticed an increase in the whole plant and root system growth of young corn plants after the application of 10 µM I of 5-ISA and 2-IBeA (2-iodobenzoic acid). Smoleń et al. [18] found that 5-ISA and 3,5-diISA, at the concentrations of 10 and 20 µM I, had no effect on the fresh weight of lettuce grown in mineral or peat soils. Other work has reported, however, no negative effect of 5-ISA, at a concentration up to 8 µM I, on the biomass of lettuce grown in the hydroponic system, while above that threshold, strong deformation of leaves and roots was observed [35]. Quite similarly, in another study with the hydroponic cultivation of lettuce, the unfavorable effect of iodosalicylates (5-ISA and 3,5-diISA) on plant growth was noted for concentrations as low as 10 µM [36]. These observations indicate a substantial impact of the method of plant cultivation on the potential toxicity of applied iodosalicylates. However, so far, no studies comparing the effect of the cultivation method on the toxicity levels of various organic iodine compounds have been conducted.
The use of organic iodine compounds as a source of that element for plant enrichment purposes is a relatively new direction in crop biofortification. Most of the so-far conducted studies were focused on the possible application of inorganic compounds, namely iodides (I − ) and iodates (IO 3 − , [10]). The present study confirmed the possibility of the uptake and translocation of exogenous iodosalicylates in dandelion plants. The highest efficiency of improving iodine content in dandelion was noted after the application of KI at a dose of 50 µM I. This stays in agreement with some of the previous reports regarding plant cultivation in pot experiments [28,37,38]. The most recognized reason for the lower uptake rate noted for IO 3 − , as compared to I − , is the required reduction of iodates prior to entrance into the plants cell [8]. The second most effective compound, in terms of increasing iodine content in dandelion plants, was 5-iodosalicylic acid. Also, Grzanka et al. [19] reported the higher efficiency of potassium iodide, followed by 5-ISA, in improving iodine levels in young corn plants. The iodine level in both leaves and roots of dandelion plants after the application of 5-ISA was higher than that after using inorganic KIO 3 , particularly at the increased dose (50 µM). Similar observations were reported by Smoleń et al. [18,35] and Sularz et al. [36]. Interestingly, the method of plant cultivation (hydroponic, pot experiment), as well as the type of substrate (peat substrate, mineral soil), had no effect on the observed relationships in lettuce plants [39]. Smoleń et al. [35] has found that the application of 5-ISA at a dose of 8.0 mM I allowed them to achieve a similar effect of iodine biofortification in lettuce leaves as KIO 3 at a dose of 40.0 mM I. The results of the present study have also allowed us to compare the efficiency of improving the iodine status of dandelion between two iodosalicylates, i.e., 5-ISA and 3,5-diISA. For low iodine doses, there were no differences regarding iodine content in dandelion plants, while the application of 50 µM 5-ISA contributed to a substantial increase in iodine accumulation in leaves and roots, as compared to 3,5-diISA. Interestingly, the level of iodine accumulation after the application of a higher dose of 3,5-diISA was similar to the effect obtained in combination with KIO 3 . That observation stays in agreement with the results presented by Smoleń et al. [39] and Sularz et al. [36]. It is worth mentioning that, apart from iodosalicylates, there also reports evaluating the applicability of other organoiodine compounds, such as iodobenzoates, in plant cultivation [34,40]. Interestingly, in the studies with corn cultivation, the effect of iodine biofortification after the application of 2-IBeA was even similar to that obtained for KI; however, the applied doses were relatively low (10 µM I [19]).
The results of 5-ISA and 3,5-diISA analysis in the leaves and roots confirmed the possibility of the uptake of iodosalicylates by dandelion plants and indicated its transport within the plant. A significant observation was that 5-ISA and 3,5-diISA accumulated more easily in roots, rather than leaves. The concentration of 5-ISA in plant leaves was approximately two-fold lower than that in roots (irrespective of the dose), while for 3,5-diISA, that ratio was approximately 1:9. On one hand, this confirms the limited but still occurring distribution of intact iodosalicylic acids within the plant. On the other, it suggests the lower mobility of 3,5-diISA in the plant.
Total iodine content was found to be higher in the combination with 5-ISA than with 3,5-diISA. However, the content of 5-ISA in plants grown in the presence of 5-ISA was lower than the content of 3,5-diISA in plants after 3,5-diISA application. This may suggest that 5-ISA, more readily than 3,5-diISA, entered and was involved in biochemical pathways resulting in its transformation into other, most likely inorganic, forms.
The content of 5-ISA in dandelion roots increased not only for the combination with 5-ISA but also in plants grown in the presence of 3,5-diISA (10 and 50 µM I), which suggests the efficient transformation of 3,5-diISA into 5-ISA in dandelion roots. However, the 5-ISA content in the leaves of plants from these combinations was not modified. On the other hand, the increased content of 3,5-diISA was noted in the roots of plants grown in the substrate enriched with 50 µM I of 5-ISA. Smoleń et al. [18] analyzed the content of iodosalicylates in the leaves, roots and root secretions of lettuce plants grown in a hydroponic system. In that study, the presence of 5-ISA was found in plants from the combinations with 5-ISA and 3,5-diISA, and the increased level of 3,5-diISA in roots was noted after the application of 5-ISA. These observations may indicate the occurrence of reverse directions of iodosalicylate transformation within a plant and requires further studies.
The total content of iodine in the control plants was lower than that after exogenous iodine was supplied. However, in both the leaves and roots of dandelion plants from that combination, the presence of low levels of 5-ISA and 3,5-diISA has been detected, which indicated that these iodosalicylates are endogenously synthesized in these plants. These results confirm previous reports on the synthesis of iodosalicylates in higher plants, without exogenous iodine application, as demonstrated for tomato [17], lettuce [18] and sweet corn [19].
One of the most important aspects of widening the popularity and applicability of biofortification approaches in agricultural practice is the maintenance or improvement of the nutritional quality of crop plants. This is also required for the works on herbal and medicinal plants, such as dandelion. One of the most studied parameters is the functioning of the antioxidant system of a plant. On one hand, it may indicate whether the introduced compound, such as iodine or iodosalicylates, acts as a stress factor; on the other hand, the increased content of respective secondary metabolites may be beneficial for the consumers. The study on the evolution of iodine functioning in marine organisms has already suggested it to be one of the first inorganic and efficient antioxidant compounds [41]. Some research has indicated that low iodine doses may also improve the tolerance of higher plants to selected stress factors, such as salinity [42] or heavy metals (Cd, [43]). Blasco et al. [44,45] widely described that the application of inorganic iodine (KI, KIO 3 ) during plant cultivation modifies the functioning of the antioxidant system on numerous levels. Numerous studies have revealed, however, that the obtained effects are dose-dependent [45][46][47]. The effect of organic iodine compounds on the antioxidant systems of higher plants has been studied to a lesser extent. Halka et al. [17,40,48] analyzed the changes in the accumulation of respective compounds (ascorbic acid, phenolic compounds), as well as the activity of antioxidant enzymes, under the influence of various iodosalicylates and iodobenzoates during the early growth of tomato plants. Sularz et al. [49] revealed that the application of 10 µM 5-ISA during lettuce cultivation had no significant effect on the contents of total phenols, as well as the antioxidant capacity of lettuce extracts, as measured using DPPH, ABTS and FRAP methods. In other work, similarly low 5-ISA doses (8 µM) did not modify the contents of phenolic compounds in lettuce leaves; however, the content of ascorbic acid-an important antioxidant molecule-decreased [18]. In the present study, two doses of iodine compounds were applied, and the contents of phenolic compounds, SA and JA, as well as the antioxidant capacity, were measured for both dandelion leaves and roots. Greater differentiation of the total contents of phenolic compounds under the influence of the type of iodine compound and doses was noted in dandelion roots, as compared to leaves. The highest root content of phenolics was noted for the combinations with KI application, which was followed by the highest values of antioxidant capacities, as measured using CUPRAC, FRAP and DPPH methods. Importantly, even though the increase in the KI dose from 10 to 50 µM caused almost a 7-fold increase in the iodine content in the roots, the difference with respect to phenolic contents and antioxidant capacity was relatively small. A significant increase in the phenolic content and antioxidant capacity of roots was also noted in the case of a higher dose (50 µM) of 3,5-diISA, even though the iodine level in that combination was lower than that after KI application. As for leaves, the highest content of phenolics was noted for the combinations with 50 µM KIO 3 , but this was not clearly related to the observed values of the antioxidant capacity. The highest antioxidant values, as measured using CUPRAC and FRAP methods, were noted in the leaves of plants treated with 3,5-diISA; however, the content of phenolics was comparable to the control value. That observation suggests that 3,5-diISA may have stimulated the synthesis of other antioxidative compounds not analyzed in the present studies. An interesting observation was a decrease in the SA content in the leaves and roots of plants with almost all combinations of iodine applications. This is consistent with the observation also noted by Halka et al. [40] and Ledwożyw-Smoleń et al. [50] in the studies on young tomato plants. In other work [51], it has been hypothesized that exogenous iodine may modulate the activity of the SAMT gene encoding S-adenosyl-L-methionine:salicylic acid carboxyl methyltransferase, which is responsible for the conversion of SA to its methyl, highly volatile form. Jasmonic acid and its derivatives are plant-signaling molecules classified into a group of phytohormones. Its level can significantly increase under stress conditions caused by biotic and abiotic factors [52,53]. In the present study, the content of jasmonic acid in dandelion leaves increased with all combinations of iodine applications, with the highest values noted for 50 µM 5-ISA and both doses of 3,5-diISA. Conversely, the root level of JA increased only after KIO 3 treatment and with all remaining combinations, including those with iodosalicylate applications, which were lower than those in the control. Unfortunately, so far, there are no available reports that would provide some further details regarding the possible interaction between iodine and jasmonic acid synthesis.

Sample Preparation
A portion of fresh leaves and roots from each combination was dried at 105 • C for 24 h in order to determine the content of dry matter. The remaining plant material was lyophilized for 48 h with the use of a Christ Alpha 1-4 freeze drier (Martin Christ Gefriertrocknungsanlagen GmbH, Osterode am Harz, Germany), ground in a laboratory grinder, sieved at 0.5 mm (FRITSCH Pulverisette 14, Idar-Oberstein, Germany) and stored in tightly closed polyethylene bags until further analysis.

Total Iodine Analysis
Total iodine content was analyzed in the lyophilized leaves and roots of dandelion plants with the use of an iCAP TQ ICP-MS + 250&1000 Gas Kits mass spectrometer (Thermo Fisher Scientific, Walthom, MA, USA) after alkaline extraction with tetramethylammonium hydroxide (TMAH; [36]). An amount of 0.1 g of lyophilized material (dandelion leaves or roots) was put into 30 mL Falcon tubes, and 10 mL of double-distilled water and 1 mL of 25% TMAH were added and mixed and incubated for 3 h at 90 • C. After cooling, samples were filled up to 30 mL with double-distilled water, mixed and centrifuged for 15 min at 4500 rpm. The supernatants were subjected to the analysis using an ICP-MS/MS spectrometer. I127 was measured in the S-SQ-KED (comprehensive interference removal with helium gas and kinetic energy discrimination) mode with Te (125Te) as an internal standard. The spectrometer was equipped with a quartz cyclonic spray chamber cooled at 2.7 • C, with a MicroMist DC Nebulizer, Ni cones and high-sensitivity interface. The operating plasma power was 1548.6 W, the nebulizer gas flow was 1.092 L/min and the helium flow in the collision cell was 4.3 L/min.

Analysis of Salicylic, Iodosalicylic and Jasmonic Acids
The contents of salicylic (SA), 5-iodosalicylic (5-SA), 3,5-diiodosalicylic (3,5-diISA) and jasmonic acids (JA) were analyzed using an LC-MS/MS system (Ultimate 3000, Thermo Scientific, Waltham, MA, USA, 4500 Q-TRAP Sciex) according to the procedure described in [36]. Here, 50 mg samples of plant material (dandelion leaves and roots) were put into 7 mL polypropylene tubes, and 5 mL of 76% ethanol with 50 ng mL −1 deuterium-labeled salicylic acid (SA-d4, Sigma-Aldrich, used as an internal standard) was added, mixed and subjected to ultrasound-assisted extraction for 1 h at 50 • C. After extraction, samples were centrifuged for 5 min at 4500 rpm. Supernatants were filtered through 0.22 µM nylon syringe filters (FilterBio NY Syringe Filter) and subjected to the analysis using LC-MS/MS. Chromatographic separation was performed using a Luna 3 µm Phenyl-Hexyl 100 Å (150 mm × 3 mm, i.d. 3 µm) column (Phenomenex, Torrance, CA, USA); the following mobile phases were applied (v:v): A-water + 0.3% formic acid; B-acetonitrile + 0.3% formic acid. The initial volume proportion of the mobile phases (60% A: 40% B) was kept for 2 min and during a further 8 min, increased linearly to reach a 98% B:2% A ratio that was kept for 4 min. After the separation, the initial mobile phase proportion (60% A: 40% B) was restored for 3 min. The injection volume was 10 µL. The mobile phase was directed to an MS ion source between 1 and 14 min of the separation. For detection, electrospray ionization (ESI) in negative ion mode was used for detection, and tandem mass spectrometry MS/MS was used for quantitative studies. Values for the precursor/product transitions of analytes are presented in Table S1; calibration curves and standard peaks for 5-ISA and 3,5-diISA are shown in Figures S1 and S2, while representative chromatographs for iodosalicylates in dandelion leaves and roots are depicted in Figure S3 (Supporting Information files). The Analyst ® 1.7 with HotFix 3 software was used for LC-MS/MS control and data processing.

Analysis of Total Phenolic Compounds
The contents of phenolic compounds, as well as the antioxidant potential of dandelion leaves and roots, were determined for 70% methanolic plant extracts. The contents of phenolic compounds in dandelion leaves and roots were analyzed after the reaction with the Folin-Ciocalteu reagent [54]. The leaf or root methanolic extract (0.25 mL) was mixed with 0.25 mL of 25% Na 2 CO 3 , 0.125 mL of the twice-diluted Folin-Ciocalteu reagent (Sigma-Aldrich, diluted twice with water prior to the analysis) and 2.25 mL of water and mixed. After a 15 min incubation in a dark place, the absorbance was measured at 760 nm (JASCO V-530 UV/Vis spectrophotometer). The final results were expressed as mg of chlorogenic acid (Sigma-Aldrich) per 1 g dry weight (chlorogenic acid equivalents, mg CAE g −1 ).
The antioxidant capacity of dandelion leaves and roots was analyzed by performing CUPRAC (cupric ion-reducing antioxidant capacity; [56]) and FRAP (ferric-reducing antioxidant power; [57]) assays using Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2carboxylic acid; Sigma-Aldrich) as an antioxidant standard. The CUPRAC method is based on the measurement of copper (II)-neocuproine reduction by the antioxidants present in the analyzed sample. Volumes of 1 mL of 10 mM CuCl 2 , 1 mL of 7.5 mM neocuproine (Sigma-Aldrich) in 96% ethanol and 1 mL of 1 M NH 4 Ac buffer, pH 7.0, were mixed with 0.3 mL of the methanolic plant extract and 0.8 mL of water. Samples were then incubated for 30 min at room temperature, and then, the absorbance was measured at 450 nm. The results were expressed as µmol Trolox per 1 g of dry matter (Trolox equivalents, µmol TE g −1 ).
The FRAP method measures the rate of the reduction of the ferric-2,4,6-Tris(2-pyridyl)s-triazine (Fe 3+ -TPTZ) complex by the antioxidants present in the sample [57]. Prior to the analysis, the FRAP working solution was prepared by mixing 300 mM acetate buffer (pH 3.6), 10 mM TPTZ (Sigma-Aldrich) in 96% ethanol and 20 mM FeCl 3 (10:1:1, v:v:v). Then, 3 mL of FRAP working solution were mixed with 0.1 mL of the plant methanolic extract and 0.3 mL of water and incubated for 30 min at room temperature. Further on, the absorbance was measured at 595 nm. The results were expressed as µmol Trolox per 1 g of dry matter (Trolox equivalents, µmol TE g −1 ).

Statistical Analysis
Statistical verification of the obtained data was conducted with the use of the ANOVA module of Statistica 13.3 software at a significance level of <0.05.

Conclusions
Plant biofortification with iodine has been proposed as one of the approaches for improving the iodine status of the human population. It seems to be of great importance particularly for those who consume a mostly plant-based diet. One of the novel directions in these studies include the usage of organoiodine compounds as iodine sources for plants, as well as evaluating the possibility of increasing the iodine content in herbal and medicinal plants. In the current study, the efficiency of the uptake, accumulation and distribution of iodine by dandelion plants was evaluated. It was confirmed that iodosalicylates (5iodosalicylic acid and 3,5-diiodosalicylic acid) are taken up and translocated within a plant. The application of 5-iodosalicylic acid during dandelion cultivation allowed to obtain the high accumulation of iodine in both the leaves and roots, even exceeding the level noted for the commonly applied inorganic iodine source, i.e., KIO 3 . It was demonstrated that after root uptake, iodosalicylates are, to a limited extent, transported into the leaves. The analysis of the plant biomass and antioxidant capacity of leaf and root extracts allows us to conclude that the applied iodine compounds and doses did not exhibit toxic effects on dandelion plants. Based on the obtained results, future studies are needed in order to characterize the metabolic processes that iodosalicylates undergo prior to or after entrance into a root cell. The interaction between iodosalicylates and the synthesis of various secondary metabolites, including those with antioxidant properties, are also yet to be discovered, particularly as they constitute the health-promoting quality of medicinal plants.