Mr Saraki and Ike Ekweremadu are accused of trying to forge the senate rules in order to help them secure their positions.
If found guilty they face up to 14 years in prison.
President Muhammadu Buhari has been at loggerheads with Mr Saraki as his party did not want him as senate leader.
Mr Buhari's preferred candidate was not present when the senate elected its president last year.
Mr Saraki's opponents say that that election was not valid and was conducted under rules which they allege were forged.
The case could have widespread political ramifications, correspondents say.
The accused arrived at court in the Nigerian capital, Abuja, in bus accompanied by senate colleagues, the BBC's Chris Ewokor reports.
Mr Saraki is also involved in another court case in which he is accused of false asset declaration, a charge he denies.