Objective To study the preparation method of Morinda officinalis iridoid glycosides (MOIG) and to explore the inhibitory effect on bone resorption of osteoclasts. Methods High-performance liquid chromatography (HPLC) was applied to determine the contents of monotropein and deacetyl asperulosidic acid from MOIG. The static adsorption-desorption test was used to screen the types of macroporous resins of AB-8, ADS-17, D101, HPD400, HPD600, HP20, S-8, SP850, XDA-1 and XDA-6, and to optimize the related parameters in process of enriching MOIG using macroporous resins. Furthermore, the osteoclasts induced from mouse bone marrow monocytes were used to evaluate the inhibitory effects of MOIG on osteoclastic bone resorption. Results After optimization, XDA-1 macroporous resin had better adsorption and desorption effects on MOIG. The optimized preparation conditions were as follows:mass concentration of MOIG in the sample solutions was 19.15 mg/mL, pH value was 1.0, diameter-height ratio of resin column was 1︰7, flow rate of loading samples was 2.0 BV/h (1 BV=80 mL), loading volume was 0.75 BV, and adsorption time was 11 h. 7 BV water was used to remove other constituents, and 10% ethanol was used to elute MOIG in the flow rate of 3.0 BV/h. The total content of monotropein and deacetyl asperulosidic acid in the prepared MOIG was more than 54%. MOIG had no significant effect on proliferation of the osteoclasts induced by mouse bone marrow monocytes, while it could significantly inhibit the tartrate-resistant acid phosphatase activity and the bone resorption of osteoclasts (P<0.05, P<0.01). Conclusion XDA-1 macroporous resin can be used to enrich MOIG, with more than 54% total content of monotropein and deacetyl asperulosidic acid. The MOIG can significantly inhibit the bone resorption of osteoclasts.