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This article presents a novel formulation for preparation of Lactobacillus casei 01 encapsulated in soy protein isolate and alginate microparticles using spray drying method. A response surface methodology was used to optimise the formulation and the central composite face-centered design was applied to study the effects of critical material attributes and process parameters on viability of the probiotic after microencapsulation and in simulated gastrointestinal conditions. Spherical microparticles were produced in high yield (64 %), narrow size distribution (d50=9.7 µm, span=0.47) and favourable mucoadhesive properties, with viability of the probiotic of 11.67, 10.05, 9.47 and 9.20 log CFU/g after microencapsulation, 3 h in simulated gastric and intestinal conditions and four-month cold storage, respectively. Fourier-transform infrared spectroscopy confirmed the probiotic stability after microencapsulation, while differential scanning calorimetry and thermogravimetry pointed to high thermal stability of the soy protein isolate-alginate microparticles with encapsulated probiotic. These favourable properties of the probiotic microparticles make them suitable for incorporation into functional food or pharmaceutical products., U radu je prikazan novi način pripreme mikrokapsula bakterije Lactobacillus casei 01 u izolatu sojinog proteina i alginata pomoću sušenja raspršivanjem. Metodologijom odzivnih površina optimiran je sastav mikrokapsula, a korištenjem plošno centriranog plana ispitan je utjecaj glavnih sastojaka kapsula te parametara procesa na preživljavanje mikroinkapsuliranog probiotika u simuliranim uvjetima gastrointestinalnog trakta. Najviše je proizvedeno kuglastih mikročestica (64 %), s raspodjelom veličina od d50=9,7 μm (raspon od 0,47) i povoljnim mukoadhezivnim svojstvima. Nakon mikroenkapsulacije preživjelo je 11,67 log CFU/g probiotika, nakon 3 sata u simuliranom želučanom soku 10,50 log CFU/g, u simuliranom soku tankog crijeva 9,47 log CFU/g, a nakon četiri mjeseca hladnog skladištenja 9,20 log CFU/g. Stabilnost probiotika nakon mikroinkapsulacije potvrđena je infracrvenom spektroskopijom s Fourierovom transformacijom, a toplinska stabilnost mikrokapsula probiotika u izolatu proteina soje i alginatu diferencijalnom pretražnom kalorimetrijom i termogravimetrijom. Zaključeno je da se zbog njihovih povoljnih svojstava mikrokapsule probiotika mogu upotrijebiti u proizvodnji funkcionalne hrane i farmaceutskih proizvoda.