Hydrolysis of food-derived opioids by dipeptidyl peptidase IV from Lactococcus lactis spp. lactis.

Food-derived opioid peptides that are released from proteins by digestion, fermentation, or food production processes lead to several health problems. The opioids are generally resistant to hydrolyze by proteases, except the dipeptidyl peptidase IV (DPPIV, EC 3.4.14.5) enzyme, because of proline amino acid. ß-casomorphin (BCM) from milk casein, gluteomorphin (GM) from wheat gluten, and soymorphin (SM) from the soybean ß-conglycinin ß-subunit are natural substrates of DPPIV because of their amino acid sequences and proline location. In the present study, DPPIV from Lactococcus lactis spp. lactis was purified and characterized by mass spectrometry. Purified DPPIV was added to standard BCM, GM, and SM, and hydrolysis percentages of morphins were measured by HPLC analysis. The results indicated that DPPIV enzyme hydrolyzed food-derived opioids (from 0.1 mM to 2 mM), BCM (33.42% for 2 mM), SM (83.81% for 2 mM), and GM (45.73% for 2 mM) in vitro. Hydrolysis percentages of SM were considerably higher than the same concentrations with BCM and GM. For dietary supplements to be promising for reducing the adverse effects of food derived opioids, this must be supported by in vivo studies of DPPIV use in the human body.

Statistical optimization of cell disruption techniques for releasing intracellular X-prolyl dipeptidyl aminopeptidase from Lactococcus lactis spp. lactis.

X-prolyl dipeptidyl aminopeptidase (PepX) is an intracellular enzyme from the Gram-positive bacterium Lactococcus lactis spp. lactis NRRL B-1821, and it has commercial importance. The objective of this study was to compare the effects of several cell disruption methods on the activity of PepX. Statistical optimization methods were performed for two cavitation methods, hydrodynamic (high-pressure homogenization) and acoustic (sonication), to determine the more appropriate disruption method. Two level factorial design (2FI), with the parameters of number of cycles and pressure, and Box-Behnken design (BBD), with the parameters of cycle, sonication time, and power, were used for the optimization of the high-pressure homogenization and sonication methods, respectively. In addition, disruption methods, consisting of lysozyme, bead milling, heat treatment, freeze-thawing, liquid nitrogen, ethylenediaminetetraacetic acid (EDTA), Triton-X, sodium dodecyl sulfate (SDS), chloroform, and antibiotics, were performed and compared with the high-pressure homogenization and sonication methods. The optimized values of high-pressure homogenization were one cycle at 130 MPa providing activity of 114.47 mU ml(-1), while sonication afforded an activity of 145.09 mU ml(-1) at 28 min with 91% power and three cycles. In conclusion, sonication was the more effective disruption method, and its optimal operation parameters were manifested for the release of intracellular enzyme from a L. lactis spp. lactis strain, which is a Gram-positive bacterium.