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Sinopodophyllum hexandrum is an important traditional Tibetan medicinal plant with great development potential in pharmaceutical industry because of the presence of major bioactive lignans such as podophyllotoxin (PT) that are widely used in combination therapy of cancer. In this study, a green and efficient method for the extraction of PT from S. hexandrum was established by optimized subcritical water extraction (SWE) combined with macroporous resin enrichment. Response surface methodology was introduced for SWE optimization to obtain the maximum yield of PT. The maximum extraction efficiency was achieved under the following conditions: extraction pressure 4 MPa, extraction solvent to feed ratio 18 mL/g, extraction temperature 180 °C, and SW flow rate 2.5 mL/min. Subsequently, an efficient macroporous resin enrichment method was developed to further purify PT in the crude extract obtained from SWE. D101 resin was selected as a proper adsorbent owing to its adsorption and desorption properties, whereas its adsorption process was found to be consistent with a pseudo-second-order model. The optimized loading amount was 3 BV of extract solution, and the desorption conditions were as follows: First, the product was eluted with deionized water until no sugar was detected in the effluent, followed by 6 BV of 20% aqueous ethanol elution and then by 15 BV of 60% aqueous ethanol elution. After the treatment with D101 resin, the concentration of PT increased from 8.3% in the crude extract to 61.5% in the final product. Moreover, the recovery yield of PT extracted with SW combined with D101 resin enrichment was 74.6%. The results show that SWE combined with macroporous resin enrichment is a green, effective, and low-cost technique for the extraction of PT from S. hexandrum. [ABSTRACT FROM AUTHOR]