This longitudinal, prospective, randomized controlled trial recruited 75 healthy pregnant women from a prenatal clinic in Taipei using convenience sampling. The participants were randomly assigned to the aromatherapy group (n=25), yoga group (n=25) or control (n=25) group using Clinstat block randomization.
The aromatherapy group received 70 minutes of aromatherapy massage once biweekly for 20 weeks; the yoga group participated in two weekly 70-minute yoga sessions led by a midwife certified as a yoga instructor for 20 weeks; the control group received only routine prenatal care. All participants' salivary cortisol and immunoglobulin A levels were collected before and after yoga or aromatherapy every 4 weeks from 16 to 36 weeks' gestation.
To collect a sufficient quantity of saliva, we used Salivette® cotton swabs (Salimetrics, State College, PA, USA), which were stored in double-layer plastic tubes with a lid. When participants chew a swab for about 2 to 3 minutes, saliva flow is stimulated to a sufficient amount (1 mL) for collection. After the tube was centrifuged at 1000 x g for 2 minutes, saliva was stored at -80°C until assay for salivary cortisol and IgA concentrations.
Cortisol was measured using a competitive enzyme-linked immunoassay (ELISA) kit (Cayman Chemical Company, USA), and salivary IgA was measured using double-antibody sandwich ELISA method per the manufacturer's instruction (ICL, Inc., USA).