## 4 Discussion  
Nowadays, an increased prevalence of malocclusions and high demands for the use of brackets have prompted researchers to assess and develop new prevention protocols for metal ion absorption from saliva while using fixed appliances (brackets). The aim of this paper was to review and evaluate the effect of fixed orthodontic appliances (brackets) on the nickel and chromium ion levels in saliva. Twelve papers were included in the systematic literature review. Seven [15; 18; 19; 20; 21; 22; 23] were clinical studies, four [2; 14; 16; 17] were in-vivo studies, and one [13] was a cross-sectional study. Synthesis of the data from the selected publications yielded controversial results. The studies summarized showed that fixed orthodontic appliances (brackets) had an impact on the levels of nickel and chromium ions in the subjects' saliva. However, the rate of increase in the ion content varied between the studies. After the placement of orthodontic appliances, the amount of nickel ions in saliva did not increase immediately in all the analyzed studies. In 2 [18; 21] out of 12 studies, the nickel content decreased after the placement of orthodontic appliances and only increased later on. The authors stated that these inaccurate data could be due to the use of different brackets produced by different manufacturers. [PERSON] et al. [18] observed that two days after the application of the brackets, nickel ion levels decreased from 153.7 ug/L to 135.3 ug/L, but started to increase a few days later; however, the nickel levels did not reach toxic levels. The WHO guideline values for drinking water currently accept a concentration of nickel of up to 70 ug/mL and a concentration of chromium of up to 50 ug/mL [14]. The authors of the paper discuss that this occurrence might be attributed to nickel binding to salivary proteins, thereby lowering its ionic form. Changes in individual salivary protein composition should therefore affect the concentration of nickel in the saliva and its absorption/distribution in the human body [7; 24]. Three studies [20] reported that saliva samples were taken only once (during treatment), which leads to debate as to whether it is worthwhile assessing saliva only during treatment without measuring it before treatment. The synthesis of the data showed that nickel and chromium ion levels tended to peak between one and six months (but did not reach toxic levels) and then gradually decreased. It is interesting to note that the results of the present report are in agreement with a previous in vitro study [25]. However, it should be noted that all the analyzed studies used different measurement units, which might also lead to disagreements in the attempts to clarify the changes in metal ion levels in saliva whilst the brackets are worn. However, in a study by [PERSON] [17] and others, a decrease in nickel concentrations was only observed after 12 months. A systematic review based on two studies [15; 20] found that mobile phone use increased the release of nickel and chromium ions and also found a positive correlation between the duration of mobile phone use and the release of nickel ions from fixed orthodontic appliances. The authors suggested that the anatomical position of the parotid salivary glands may have an influence on these changes in ion content [15]. The studies evaluated in the publications were carried out in a standardized way: saliva samples were collected using a standardized method, after rinsing the mouth with distilled water, and then the unstimulated saliva was placed in a saliva collection container. The saliva collection method can be stimulated or unstimulated. In all the studies, the saliva sample was collected without stimulation, as in this case, about two thirds of the total amount of the saliva is secreted by the sublingual salivary glands.