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Various opinions have been suggested regarding non suicidal self-injury (NSSI) and pain relationship. Opiorphin is a recently found peptide that inhibits enkephaline-catabolizing enzymes. Analgesia caused by opiorphine has been demonstrated in animal experiments. No studies have examined the relationship between opiorphin and pain sensation until today. We aimed to investigate opiorphine and pain threshold among self-injuring adolescents. Adolescents aged 14-18 years were included in the study. The NSSI group consisted of 37 adolescents diagnosed with NSSI according to DSM-V Section 3, and the non-NSSI group consisted of 36 adolescents without any psychiatric disorder. We measured pain threshold with a pressure sensitive algometer device and analyzed saliva opiorphin levels by ELISA method. Mediation analysis was performed using Process Macro developed by Hayes. NSSI group had statistically significantly higher pain threshold and opiorphin levels than the non-NSSI group. There was a positive correlation between pain threshold values and opiorphin levels in the NSSI group. Also, a positive correlation between opiorphin levels and total cutting episode number was found. We searched for a probable relationship of pain threshold with episode number of each type of NSSI act. Accordingly, a positive correlation with two major act and a negative correlation with two minor act was shown. The opiorphine was found to be a mediator variable in the relationship between the pain threshold and the cutting number. The relationship between opiorphin, pain threshold and cutting number and their mediating effects with each other may highlight the pain-based biological origins of NSSI.