Makola is unlikely to feature in Saturday night's last-32 tie at Orlando Stadium due to a back problem.
Fellow midfielder Musa Nyatama is suspended for the contest, while the versatile Abbubaker Mobara is still undergoing rehabilitation with the club's strength and conditioning team.
Thabo Qalinge, Marc van Heerden and Gladwin Shitolo are also nursing knocks.
Pirates released a statement on their website, part of which reads: 'Mpho Makola is doubtful for the cup clash tomorrow as he continues to receive treatment on a back injury.
'According to the Bucs medical team, it’s unlikely that the 31-year old will recover in time for the match.
'The absence of both Nyatama and Makola is exacerbated by the injury to Abbubaker Mobara who is also still undergoing rehabilitation with the strength and conditioning team.
'Fortunately for the Pirates technical team, newly signed midfielder Xola Mlambo is available for selection amongst others.'
Mlambo made his Bucs debut off the bench in a 2-1 win over Maritzburg United last weekend following his signing from Bidvest Wits.