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Sajid Javid is under pressure for still holding share options in a tech firm that provides artificial intelligence (AI) software to the health sector.
The health secretary was paid more than £150,000 a year by C3.ai, a California firm specialising in AI, between October 2020 and June 2021, when he rejoined the cabinet in his current role.
The party's deputy leader Angela Rayner has written to the prime minister's ethics adviser Lord Geidt alleging it is a "clear conflict of interest and breach of the ministerial code".
Reports suggest the options are worth around £45,000.
Mr Javid is said to have begun the process of divesting after taking the job of health secretary in the summer.
"In September, the secretary of state's department announced that the use of 'artificial intelligence' would shorten waiting lists in our NHS," Ms Rayner's letter reads.
"The Department of Health and Social Care announcing that it is going to spend taxpayers' money on artificial intelligence could clearly be perceived as beneficial to an artificial intelligence company, and to the value of shares in an artificial intelligence company such as C3.ai."
Labour's deputy leader then references the section of the ministerial code that states ministers "must scrupulously avoid any danger of an actual or perceived conflict of interest" between their role and their private financial interest.
A spokeswoman for the Department of Health and Social Care said: "The secretary of state has acted in line with the ministerial code and has properly declared these share options in the usual way."
Meanwhile, a source close to Mr Javid told the PA news agency that the former chancellor started the process of divesting the shares when he re-entered the cabinet - but could not say when the process would be complete.
The letter comes as MPs face intensified scrutiny over their roles outside parliament following a sleaze row which reared its head over the case of Owen Paterson.
The former Tory MP resigned after being found to have carried out an "egregious" breach of lobbying rules.
Mr Javid became health secretary in June when Matt Hancock resigned after being caught breaking coronavirus restrictions on camera, kissing an aide in his departmental office.
Sky News has contacted C3.ai for comment.