Around 180,000 passengers will be affected by the walkout, Lufthansa said on Monday, after last-minute talks with pilots' union Vereinigung Cockpit failed to reach a wages deal.
A spokesman for the group told AFP it would seek an injunction from a Munich labour court to block the strike, although a similar request was rejected by a Frankfurt court last week and the airline lost a second time at appeal.
Pilots have been battling the German behemoth since April 2014 over pay.
During a two-day pause in the strike on Sunday and Monday, Lufthansa offered the flight crew a 4.4-percent pay rise spread over two years, as well as a one-off payment equivalent to two months' wages.
But pilots stuck to their demand of a pay rise of an average of 3.66 percent per year, retroactive for the past five years.
“Unfortunately, high-level talks held today at short notice failed to lead to an agreement on the wage contract,” said Jörg Handwerg, board member at Cockpit, on Sunday.
“It is completely incomprehensible that (Lufthansa) has refused to put forward an offer that can at least form the basis of a negotiation,” he added.
The latest round of industrial action, which started on Wednesday, is the 14th since April 2014.
While Tuesday's walkout will only affect short-haul flights from Germany, Wednesday will see some long-haul services also affected.
Lufthansa said the flight plan for Tuesday and Wednesday would bring the number of passengers affected over the week of strikes to more than 525,000, with 4,461 cancelled flights.
While it strives to slim down and fend off competition from low-cost rivals like Easyjet and Ryanair, Lufthansa has been snarled in long-running disputes with pilots and cabin crew over the past two years.
The airline reached a pay and conditions deal with cabin crew in July after they staged the firm's longest-ever walkout in November 2015.
That seven-day stoppage saw 4,700 flights cancelled and more than half a million passengers grounded.
The deal with cabin crew also included a no-strike agreement until 2021 in exchange for job guarantees.
Lufthansa in October said it expected annual earnings before interest and tax in 2016 to reach “approximately the same level” as 2015's €1.8 billion ($1.9 billion) figure.