Interlayer adhesion is one of the most pivotal aspects related to mechanical performance and durability of 3D printed concrete. However, due to continuous printing of multi-layered structures, large-scale buildings require long time intervals between layer deposition, with interlayer bond strength decreasing as time intervals between printed layers increase. This study evaluates comprehensively the effects of time intervals between layers: 1 h, 1.5 h, 2 h, 3 h, and 4.5 h as well as two different height to span ratios (hb/s). In order to assess the properties of interlayer regions microstructural examinations using techniques such as X-ray micro-computed tomography (micro-CT), scanning electron microscopy (SEM) were applied. In addition destructive testing of compressive, splitting tensile and flexural strengths along with crack opening analysis using the Digital Image Correlation (DIC) method was performed and supported with FEM analysis. The results show that interlayer adhesion strength decreased significantly for time intervals of up to 2 h, with a linearly decreasing trend. The reduction in strength the interlayer bond for a 2-h interval was 56.6%, compared to the reference specimen. Afterwards, the decrease in strength was minimal (up to 68.5% after 4.5 h time interval). However, microstructural investigations revealed that specimens with printing interval beyond 2 h exhibited noticeable continuous cracking in interfacial zone with widths reaching of up to 100 μm (SEM). At the same time micro-CT analysis confirmed creation of longitudinal pores which in turn was reflected in different cracking phenomena of specimens with longer printing intervals (>2 h). This phenomenon was further explained through FEM modeling. As an outcome it was proven through DIC that that increasing the time interval between the deposit of successive layers not only affects the strength of the entire beam, but also affects its cracking mode. The discontinuities in the interlayer contact zone leads to destruction characteristic of composite structures, thus the structure comes to be destroyed layer by layer, resulting from insufficient interlayer adhesion. The presented study is the first to fully evaluate the failure behavior of 3D printed specimens with long time intervals.