Background Jejunal anastomosis in horses with one- or two-layer conventional technique can lead to leaks at the anastomosis site and the possibility of severe septic peritonitis. The objective of this study was to compare the use of a recently designed UV-polymerizable methacrylate adhesive as the second layer of a two-layer anastomosis with a Cushing pattern.Fifteen fresh harvested jejunum segments were collected from horses euthanized for unrelated reasons and owner consent was obtained for research donation. Each segment was divided in 3 pieces, each assigned to 3 different groups. In 2L-CT group, resection and anastomosis was performed using a double-layer simple continuous/Cushing suture. In 1L-UV-PMA group, resection and anastomosis was performed using a single-layer continuous technique sealed with a UV-polymerizable methacrylate adhesive. Control group was left untouched. Anastomotic construction time of the second layer, Bursting Strength Pressure (BSP), Luminal Diameter Reduction (LDR), and mode of failure were measured and compared between groups.Results The construction time (Mean [95% CI]) was 3.02 min [2.50; 3.55] in 1L-UV-PMA group and 8.09 min [7.59; 8.61] in 2L-CT group. The difference was significant (P<0.001).The BSP (Mean [95% CI]) was 170.47 mmHg [146.29; 194.65] in 1L-UV-PMA group, 175.33 mmHg [156.83; 193.83] in 2L-CT group, and 189.93 mmHg [162.52; 217.34] in the control group. The difference was significant only between the 1L-UV-PMA group and the control group (P=0.04).The LDR (Mean [95% CI]) was 51% [47; 55] in 1L-UV-PMA group and 48% [43; 53] in 2L-CT group. The difference was not significant (P=0.26).Eight segments ruptured on the suture line in the 1L-UV-PMA group and six segments ruptured on the suture line in the 2L-CT group. The difference was not significant (P=0.36).On macroscopic evaluation, the 1L-UV-PMA anastomosis formed a tunnel-like anastomosis. After testing, some of the samples from the 1L-UV-PMA group showed shreds of glue detached from the serosa.Conclusions Complete covering of anastomosis with UV-PMA glue is comparable in terms of leaking pressure and Luminal Diameter Reduction but faster to perform than an inverting suture pattern. Modification of the technique is warranted to decrease tunneling at the anastomosis site.