The invention discloses a sulfur-containing thick and mixed tetracyclic azasugar derivative, which has a general chemical formula as shown in figure I. A preparation method of the sulfur-containing thick and mixed tetracyclic azasugar derivative comprises the following steps of (a) mixing pentabasic azasugar aldehyde and amino-acid ester hydrochloride, dissolving in an organic solvent, stirring for 30-60min at the temperature of 40-60 DEG C, adding sulfydryl salicylic acid or sulfydryl nicotinic acid, stirring, reacting for 8-15h at the temperature of 40-100 DEG C, adjusting a pH value of reaction liquid to be neutral, extracting, and obtaining an intermediate compound as shown in a general chemical formula IV; (b) removing a protecting group from the intermediate compound IV under the acidic condition, amidating ring closure under the alkaline condition, and obtaining the sulfur-containing thick and mixed tetracyclic azasugar derivative as show in the general chemical formula I. The preparation method of the sulfur-containing thick and mixed tetracyclic azasugar derivative provided by the invention is simple, easy to operate, and easy to produce in large scale; through experimental verification, the prepared compound I has stronger HIV (Human Immunodeficiency Virus) inhibitory activity, and can be popularized and applied in anti-HIV medicinal preparation.