Ancient grains have gained considerable attention in recent years, as some research suggests they may be healthier than modern wheat. The present study aims to evaluate the chemical, rheological, and microbiological features of three Southern Italian cultivated ancient wheat varieties: Risciola, Carosella, and Saragolla. ATR-FTIR analyses were performed on the finely ground grain samples of the three varieties. The selected grains were ground with a stone mill, and different sifting degrees (whole—100%, type 1—80%, and type 0—72%) were evaluated. The flours showed a good nutritional profile, a higher amylose/amylopectin ratio, and a lower glycemic index than the literature. The gluten index of the samples was in the range 2.6–28.9%, and the flours can be classified as weak, having a value <30%. The farinographic test showed a short development time, low dough stability, a high softening degree, and water absorption, which increased with the degree of sifting. Microbiological analyses performed on flours from ancient grains at different degrees of sifting show their safety, according to their microbiological parameters, which fall within the legal microbiological requirements established by the European Commission Regulation (EC).