Alistair Carmichael has accused the Scottish Government of using "heavy influence" to silence business voices north of the Border that are pro-UK.
He says firms are reluctant to enter into the independence debate for fear of being punished.
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In an interview with The Herald to be published in full tomorrow, the Secretary of State for Scotland also rebuts the notion that Prime Minister David Cameron is semi-detached in the referendum campaign, declaring: "I don't accept the assertion that he is a backseat driver."
Pro-UK politicians have expressed frustration that business leaders have felt unable to air their concerns about independence in public.
Earlier this month, the boss of a FTSE 100 company reportedly told Scottish Government representatives he could not make investment decisions in Scotland because of the uncertainty about the tax regime and claimed "the mood of the meeting immediately became very dark; they became very aggressive".
Mr Carmichael said: "We hear increasingly about people, business voices in particular, being disinclined to enter the debate because they think they will be punished as a result."
Asked if he was saying they were being intimidated by the Scottish Government to keep quiet, he replied: "Intimidation is a strong word but there is certainly heavy influence. And you can imagine a situation as the campaign progresses the heaviness of that influence does become inappropriate."
Mr Carmichael said the experience of the FTSE 100 boss was not an isolated incident. "I hear of it time and again. It's always told to me in terms, 'But we don't want you to talk about this publicly'. So people are not yet willing to stick their head above the parapet."
Asked if there was a fear factor, Mr Carmichael replied: "There is very much. This is what business people tell me; that they are scared of the consequences for their business of getting on the wrong side of the Scottish Government."
Without naming names, he noted: "I see it in the media sector. I have seen media outlets feel constrained in their coverage."
Yet he said Scotland's captains of industry would not be able to keep their "grumblings" over independence quiet for much longer as the campaign during 2014 intensified
However, a spokesman for Scottish Finance Secretary John Swinney insisted Mr Carmichael's claim of "heavy influence" by the Scottish Government was not true and that ministers in Edinburgh welcomed "all voices and views to the debate on Scotland's future".
The spokesman said: "These claims are wrong. We welcome all voices and views to the debate on Scotland's future; and no one familiar with this debate can seriously suggest it is anything other than open and energetic with full, frank and vigorous contributions from all sides."
He pointed to a recent poll from the British Chambers Of Commerce, which concluded the referendum and the prospect of an independent Scotland had 'left most businesses unfazed'.
"The reality is businesses are investing heavily in Scotland in the run-up to the referendum, with the latest survey report on inward investment from Ernst and Young showing that inward investment projects are coming to Scotland at their highest level for 15 years," he added.
Also in the interview, Mr Carmichael brushed aside the idea that Mr Cameron was taking a low profile in the campaign for fear of stirring harmful anti-Conservative sentiment among Scottish voters.
He said: "I don't accept the assertion that he is a backseat driver. He is the Prime Minister of the whole of the United Kingdom. He spends time in Scotland both personally and politically. He is completely engaged in the debate, I know because I speak to him about it."
Meantime, Deputy Prime Minister Nick Clegg yesterday described the Scottish Government's proposals on currency union as "rather erratic and opaque". He said he put the "huge and possibly economically crippling uncertainty" around the currency in the same category as the "very unclear scenario" regarding an independent Scotland's status in the EU.