It has lodged a formal objection with the European Commission (EC) in a bid to overturn the plan, one week after Bulgaria became the first EU state to raise objections.
The EC extended the consulting period on the minimum price to the end of the year, causing a possible delay to the Scottish Government's plan to introduce it as early as next April.
Poland and Austria have also signalled they are considering the issue and it is believed Spain and Portugal are preparing objections.
MSPs passed legislation paving the way for minimum alcohol price last May, but the Scottish Government agreed to notify European countries whose drinks industries might be affected.
Under Government proposals to set the price at 50p per unit of alcohol, it will become illegal to sell a standard bottle of wine for less than £4.69 or a bottle of whisky for less than £14.
The objections are based on laws designed to protect free trade among EU member states. Countries able to produce relatively cheap wine or spirits have argued the minimum price will destroy their competitive advantage.
Europe-wide trade bodies representing wine and spirits producers have already lodged objections.
Miles Beale, chief executive of the Wine and Spirit Trade Association, said: "Opposition to the Scottish Government's minimum unit pricing plans is growing in Europe.
"The UK Government will now be required to explain to the European Commission the legality of the Scottish Government's proposals. The legality of the proposals have always been questioned and it now seems that there is a groundswell of opposition to minimum unit pricing building from EU member states."
The Scottish Government said it had expected objections but remained confident it would be able to justify the policy on health grounds.
Ministers claim alcohol-related deaths will fall by 60 in the first year of the policy and, after 10 years, it will save more than 300 lives annually.
Crime is expected to be cut by 3500 offences per year.
A Scottish Government spokesman said: "The Scottish Government believes minimum pricing will save lives and reduce the harm caused by alcohol misuse, we also believe minimum unit pricing is the most effective pricing measure.
"We are confident that we can demonstrate that the minimum pricing of alcohol is justified in Scotland on the basis of public health and social grounds."
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