Objective: The objective of this study was to design a method to identify patients at risk for preterm premature rupture of the membranes using a simple assay of salivary proteinase activity. Study design: Saliva samples were collected from women in the following groups using Salivette: (1) nonpregnant control; (2) during the second trimester of pregnancy; (3) during active labor at term; (4) women with premature rupture of the membranes before preterm delivery; and (5) postpartum (within 3 hours after delivery at term). Total proteolytic activity in saliva samples was measured by fluorometry using the generic substrate DQ-gelatin in the presence of specific inhibitors to selectively detect matrix metalloproteinase activities. The concentrations of various matrix metalloproteinases in saliva samples were also measured by multiplex bead assay using the Luminex platform. Results: All saliva samples exhibited detectable matrix metalloproteinase activity. Salivary matrix metalloproteinase activity is low in nonpregnant females (0.27 ± 0.15) and increases in samples taken in the second trimester (0.5 ± 0.5) and at term during active labor (1.03 ± 1.2). Samples collected from women with premature rupture of the membranes before preterm delivery had the highest activity (2.5 ± 3.7) followed by postpartum after normal term delivery (2.1 ± 1.6). The matrix metalloproteinase activity was higher in premature rupture of the membranes before preterm delivery samples, compared with all other stages of pregnancy. Multiplexmatrix metalloproteinase assay documented a significant increase in total matrix metalloproteinase- 9 concentration in saliva from premature rupture of the membranes before preterm delivery, compared with any of the other groups. Similarly matrix metalloproteinase-9 activity was also significantly increased in premature rupture of the membranes before preterm delivery group, compared with all others. Conclusion: Herein we report a simple test to monitor proteolytic enzyme activity in saliva during pregnancy. The highest matrix metalloproteinase activity is seen in premature rupture of the membranes before preterm delivery and postpartum samples. Ongoing studies aim to further define salivary proteinase activity in patients at high risk for premature rupture of the membranes before preterm delivery and to evaluate its potential as a predictive test for premature rupture of the membranes before preterm delivery. Objective: The objective of this study was to design a method to identify patients at risk for preterm premature rupture of the membranes using a simple assay of salivary proteinase activity. Study design: Saliva samples were collected from women in the following groups using Salivette: ( (2) during the second trimester of pregnancy; (3) during active labor at term; (4) women with premature rupture of the membranes before preterm delivery; and (5) postpartum (within 3 hours after delivery at term). Total proteolytic activity in saliva samples was measured by fluorometry using the generic substrate DQ-gelatin in the presence of specific inhibitors to selectively detect matrix metalloproteinase activities. The concentrations of various matrix metalloproteinases in saliva samples also also measured by multiplex bead assay using the Luminex platform. Results: All saliva samples absorbed detectable matrix metalloproteinase activity. Salivary matrix metalloproteinase The activity is low in nonpregnant females (0.27 ± 0.15) and increases in samples taken in the second trimester (0.5 ± 0.5) and at term during active labor (1.03 ± 1.2). Samples collected from women with premature rupture of the membranes before preterm delivery had the highest activity (2.5 ± 3.7) followed by postpartum after normal term delivery (2.1 ± 1.6). The matrix metalloproteinase activity was higher in premature rupture of the membranes before preterm delivery samples, compared with all other stages of pregnancy. Multiplexmatrix metalloproteinase assay documented a significant increase in total matrix metalloproteinase- 9 concentration in saliva from premature rupture of the membranes before preterm delivery, compared with any of the other groups. group, compared with all others. Conclusion: Herein we report a simple test to monitor prot eolytic enzyme activity in saliva during pregnancy. The highest matrix metalloproteinase activity is seen in premature rupture of the membranes before preterm delivery and postpartum samples. Ongoing studies aim to further define salivary proteinase activity in patients at high risk for premature rupture of the membranes before preterm delivery and to evaluate its potential as a predictive test for premature rupture of the membranes before preterm delivery.