The aim of this study is to investigate the effect of methylphenidate treatment on oxidative stress by measuring the levels of Total Oxidant Status (TOS), Total Antioxidant Status (TAS), Oxidative Stress Index (OSI=TOS/TAS), Malondialdehyde (MDA), Oxidized LDL (Ox-LDL), and Superoxide Dismutase (SOD) in serum samples of patients diagnosed with Attention Deficit Hyperactivity Disorder (ADHD) who have either started or are planned to start methylphenidate treatment. The measurements will be taken at baseline (0 months, before treatment initiation) and at the 3rd month (after treatment has begun) to assess the effect of methylphenidate on oxidative stress. Patients who were already planned to begin methylphenidate treatment will be invited to participate in this study. Since this is not an interventional study, no additional treatments will be administered or altered beyond the treatment the patient is already required to receive.
The study population will consist of patients who are diagnosed with "Attention Deficit Hyperactivity Disorder (ADHD)" according to DSM-5-TR, who have applied to the child and adolescent psychiatry outpatient clinics of Bakırköy Prof. Dr. Mazhar Osman Mental Health and Neurological Diseases Training and Research Hospital and have been started or planned to start methylphenidate treatment by the physician they have been examined by. After an in-clinic announcement, these patients will be directed to the researcher, Enes Faruk Altunkılıç. After being informed both verbally and in writing through the informed consent process, those who agree to participate, sign the informed consent form, and meet the inclusion and exclusion criteria will be included in the study. It will also be explained to the patients and their families that their participation or non-participation in this study will not affect the treatment they will receive. After informed consent, a structured clinical interview for DSM-5-TR will be conducted using the "Mood Disorders and Schizophrenia Form for School-Aged Children - Now and Lifetime DSM-5 - Turkish Adaptation (ÇDŞG-ŞY-DSM-5-T)." To obtain sociodemographic and clinical data for the participants, the "Sociodemographic and Clinical Data Form" created by the researchers will be completed. For the ADHD diagnosed group, the "Conners Parent Rating Scale - Revised Short" and "Conners Teacher Rating Scale - Revised Short" will be applied to determine the severity of the disorder, symptoms, and predominant subtyping. After the diagnosis is made and evaluated according to the exclusion criteria, blood samples will be taken from the patient after a 10-12 hour fasting period, between 9-12 AM, in a yellow-capped tube, before the routine methylphenidate treatment is used. After waiting for 10-20 minutes at room temperature, the sample will be centrifuged at 3000 RPM for 20 minutes, and the serum will be collected in Eppendorf tubes and stored at -80°C until analysis. After 3 months of treatment, blood samples will be taken again in the same manner and stored. Once all the samples are collected, the serum samples will be analyzed for total antioxidant status (TAS), total oxidant status (TOS), malondialdehyde (MDA), superoxide dismutase (SOD), and oxidized LDL levels according to the human ELISA kit protocols at the Biochemistry Laboratory of Bakırköy Dr. Sadi Konuk Training and Research Hospital by biochemist Dr. Hacer Eroğlu İçli. After 3 months of treatment, the Conners Parent Rating Scale - Revised Short and Conners Teacher Rating Scale - Revised Short forms will be applied again. In addition to the markers, the Oxidative Stress Index (OSI = TOS/TAS) will be calculated and included in the evaluation before and 3 months after the treatment.
Study Type
OBSERVATIONAL
Enrollment
39
This is not an interventional study. Patients diagnosed with Attention Deficit Hyperactivity Disorder (ADHD), for whom methylphenidate treatment is planned, have been invited to participate in the study. The patients have been enrolled in follow-up, and our contact information has been provided to them. As part of routine care, patients have been called in once a month. Necessary dosage adjustments have been made in accordance with medical guidelines, without any intervention for the study. Patients with complaints or those who needed to be seen earlier have been seen in between. As a result, the patients were followed for a total of 3 months for this study. Serum samples for oxidative stress markers were collected at baseline (before treatment started) and at the 3rd month (after treatment began).
Bakırköy Prof. Dr. Mazhar Osman Mental Health and Neurological Diseases Training and Research Hospital
Istanbul, Bakirkoy, Turkey (Türkiye)
Change in Total Oxidant Status
Total Oxidant Status (TOS) level was measured according to the human ELISA kit protocol. Serum samples from 39 patients were compared immediately before and after 3 months of treatment. TOS was measured according to the human ELISA kit protocol as follows (bioassay technology laboratory, Cat.No E1599Hu): Standards added to wells. 40μl of serum sample was added to the sample wells, followed by 10μl of anti-TOS antibody. Incubated at 37°C for 60 minutes. plate was washed 5 times with wash buffer. 50μl of substrate solution A was added to the wells and then 50μl of substrate solution B was added to each well. The plate was incubated for 10 minutes at 37°C in the dark. 50μl of Stop Solution was added to the wells and the blue color immediately changed to yellow. The optical density (OD value) of each well was determined using a microplate reader set to 450 nm within 10 min after adding the stop solution.
Time frame: Immediately before starting the treatment and up to the 3rd month of treatment.
Change in Total Antioxidant Status
Total antioxidant status (TAS) level was measured according to the human ELISA kit protocol. Serum samples from 39 patients were compared immediately before and after 3 months of treatment.TAS was measured according to the human ELISA kit protocol as follows (bioassay technology laboratory, Cat.No E4350Hu): Standards added to wells. 40μl of serum sample was added to the sample wells, followed by 10μl of anti-TOS antibody. Incubated at 37°C for 60 minutes. plate was washed 5 times with wash buffer. 50μl of substrate solution A was added to the wells and then 50μl of substrate solution B was added to each well. The plate was incubated for 10 minutes at 37°C in the dark. 50μl of Stop Solution was added to the wells and the blue color immediately changed to yellow. The optical density (OD value) of each well was determined using a microplate reader set to 450 nm within 10 min after adding the stop solution.
Time frame: Immediately before starting the treatment and up to the 3rd month of treatment.
Change in Superoxide Dismutas
superoxide dismutas level was measured according to the human ELISA kit protocol. Serum samples from 39 patients were compared immediately before and after 3 months of treatment. Superoxide dismutas was measured according to the human ELISA kit protocol as follows (bioassay technology laboratory, Cat.No E0918Hu): Standards added to wells. 40μl of serum sample was added to the sample wells, followed by 10μl of anti-TOS antibody. Incubated at 37°C for 60 minutes. plate was washed 5 times with wash buffer. 50μl of substrate solution A was added to the wells and then 50μl of substrate solution B was added to each well. The plate was incubated for 10 minutes at 37°C in the dark. 50μl of Stop Solution was added to the wells and the blue color immediately changed to yellow. The optical density (OD value) of each well was determined using a microplate reader set to 450 nm within 10 min after adding the stop solution.
Time frame: Immediately before starting the treatment and up to the 3rd month of treatment.
Change in Malondialdehyde
Malondialdehyde (MDA) level was measured according to the human ELISA kit protocol. Serum samples from 39 patients were compared immediately before and after 3 months of treatment. MDA was measured according to the human ELISA kit protocol as follows (bioassay technology laboratory, Cat.No E1371Hu): Standards added to wells. 40μl of serum sample was added to the sample wells, followed by 10μl of anti-TOS antibody. Incubated at 37°C for 60 minutes. plate was washed 5 times with wash buffer. 50μl of substrate solution A was added to the wells and then 50μl of substrate solution B was added to each well. The plate was incubated for 10 minutes at 37°C in the dark. 50μl of Stop Solution was added to the wells and the blue color immediately changed to yellow. The optical density (OD value) of each well was determined using a microplate reader set to 450 nm within 10 min after adding the stop solution.
Time frame: Immediately before starting the treatment and up to the 3rd month of treatment.
Change in Ox-LDL
Ox-LDL level was measured according to the human ELISA kit protocol. Serum samples from 39 patients were compared immediately before and after 3 months of treatment. Ox-LDL was measured according to the human ELISA kit protocol as follows (bioassay technology laboratory, Cat.No E1521Hu): Standards added to wells. 40μl of serum sample was added to the sample wells, followed by 10μl of anti-TOS antibody. Incubated at 37°C for 60 minutes. plate was washed 5 times with wash buffer. 50μl of substrate solution A was added to the wells and then 50μl of substrate solution B was added to each well. The plate was incubated for 10 minutes at 37°C in the dark. 50μl of Stop Solution was added to the wells and the blue color immediately changed to yellow. The optical density (OD value) of each well was determined using a microplate reader set to 450 nm within 10 min after adding the stop solution.
Time frame: Immediately before starting the treatment and up to the 3rd month of treatment.
Change in Oxidative Stress Index (OSI)
Oxidative Stress Index (OSI) level was measured according to the human ELISA kit protocol. Serum samples from 39 patients were compared immediately before and after 3 months of treatment. OSI is an index created by the ratio of TOS to TAS. OSI is a parameter that shows the direction in which the body's oxidative stress balance shifts. An increase or decrease in the OSI value indicates that the oxidative balance has changed. While an increase in OSI value indicates a change in balance in favor of oxidants; a decrease in OSI value indicates a change in balance in favor of anti-oxidants.
Time frame: Immediately before starting the treatment and up to the 3rd month of treatment.
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