Simultaneous quantification of buprenorphine, naloxone and phase I and II metabolites in plasma and breastmilk by liquid chromatography-tandem mass spectrometry

J Chromatogr A. 2016 May 13:1446:70-7. doi: 10.1016/j.chroma.2016.03.076. Epub 2016 Mar 31.


Opioid abuse during pregnancy is associated with fetal growth restriction, placental abruption, preterm labor, fetal death, and Neonatal Abstinence Syndrome. Current guidelines for medication-assisted opioid addiction treatment during pregnancy are methadone or buprenorphine monotherapy. Buprenorphine/naloxone combination therapy (Suboxone(®)) has not been thoroughly evaluated during pregnancy and insufficient naloxone safety data exist. While methadone- and buprenorphine-treated mothers are encouraged to breastfeed, no studies to date investigated naloxone concentrations during breastfeeding following Suboxone administration. For this reason, we developed and fully validated a liquid chromatography-tandem mass spectrometry method for the simultaneous quantification of buprenorphine, buprenorphine-glucuronide, norbuprenorphine, norbuprenorphine-glucuronide, naloxone, naloxone-glucuronide and naloxone-N-oxide in 100μL human plasma and breastmilk in a single injection following protein precipitation and solid-phase extraction. Lowest limits of quantification were 0.1-2μg/L with 20-100μg/L upper limits of linearity. Bias and imprecision were <±16%. Matrix effects ranged from -57.9 to 11.2 and -84.6 to 29.3% in plasma and breastmilk, respectively. All analytes were stable (within ±20% change from baseline) under all tested conditions (24h room temperature, 72h at 4°C, 3 freeze/thaw cycles at -20°C, and in the autosampler for 72h at 4°C). For proof of concept, buprenorphine and its metabolites were successfully quantified in authentic positive maternal and infant plasma and paired breastmilk specimens. This comprehensive, highly sensitive and specific method detects multiple buprenorphine markers in a small specimen volume.

Keywords: Breastmilk; Buprenorphine; Liquid chromatography–tandem mass spectrometry; Naloxone; Plasma; Prenatal drug exposure.

MeSH terms

  • Buprenorphine / analogs & derivatives*
  • Buprenorphine / analysis
  • Buprenorphine / blood
  • Chromatography, Liquid
  • Female
  • Glucuronides / analysis*
  • Glucuronides / blood
  • Humans
  • Infant, Newborn
  • Metabolic Detoxication, Phase I
  • Metabolic Detoxication, Phase II
  • Methadone / analysis
  • Milk, Human / chemistry*
  • Naloxone / analysis*
  • Naloxone / blood
  • Narcotic Antagonists / analysis*
  • Narcotic Antagonists / blood
  • Pregnancy
  • Solid Phase Extraction
  • Tandem Mass Spectrometry


  • Glucuronides
  • Narcotic Antagonists
  • Naloxone
  • Buprenorphine
  • norbuprenorphine
  • Methadone