Win32::API::Prototype and Win32::API::Import effectively do the same thing. They both export a sub into the callers name space that does:

sub ApiName {
    return $globalHash{ 'ApiName' }->Call( @_ );
}
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The name of the global hash is different between the two packages, but the performance should be close to identical.

You will get a minor performance gain by avoiding the import sub. Ie:

usw Win32::API;

my $api = Win32::API->new( 'dll', 'api', 'II', 'I' ) or die $^E;

$api->Call( @_ );
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You avoid an extra level of perl subroutine call and a hash lookup. For all but the lowest cost APIs, the difference will be minimal, as most of the cost is in converting the call parameters to their C equivalents before calling into the DLL. It might make some perceptible difference for calls that take no perameters.

If performance is a realistic consideration, you'd be better off writing Inline::C wrappers for the APIs you need.

but my prog. takes lot of time in win-api function call

The difference between the two is the declaration of the API function, which happens one time in your script (if you're doing it right). The difference between should be no more than, say, 100µSec. At worst.

The calls to the function should be (close to) identical under the hood, so they should take virtually the same time.

Seeing how Win32::API::Prototype is based on Win32::API, I don't expect there to be any time difference