Leader of Opposition Ramesh Chennithala said the entry of the women into the shrine "hurt" the sentiments of devotees and it shows the headstrong attitude of the chief minister to take women of the traditionally "barred" age group to the temple.
A senior temple official told the BBC that the sanctum has now reopened.
SKS activists vowed not to rest till Vijayan was ousted from office and called a dawn-to-dusk shutdown on Thursday.
The workers hurled stones at the women and policemen, injuring three personnel, police said, adding that they had to fire in the air and burst teargas shells to disperse the attackers.
"The government had assured us all help. Barring minor protests, there was no other issue".
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"CPI (M) agenda is clear now".
Expressing solidarity, thousands of men also lined up parallel forming a second human "wall".
However, the Vyapari Vyavasayi Ekopana Samithi (VVES), a Kerala-wide union of traders and merchants, has said their shops would not be shut on Thursday.
The Sabarimala shrine was historically closed to women of "menstruating age" - defined as between 10 and 50.
He said, "Unique tantric traditions and ritualistic practices seldom permit menstruating women to the Ayyappa Temple at Sabarimala". However, India's Supreme Court struck down the ban in September, saying that it violated gender equality laws.
"'Being the chief priest, I am duty-bound to follow the ritualistic practices at Sabarimala", he said. "The government only did its role in upholding the Supreme Court directive", he told journalists. Some protesters can also be seen at the spot in the video.
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The two women in their forties entered the temple early Wednesday and worshipped there, said Pramod Kumar, the state police spokesman. "The role of the government is to abide by the Supreme Court order and it has done just that".
Police have provided security to the homes of the two women fearing possible protests.
Protests soon erupted in the state with BJP Mahila Morcha members protesting outside the state secretariat.
"What they did was to act like cowards".
The Supreme Court on September 28 overruled a decades-old ban on women of menstruating age at Sabarimala, which is a four-hour uphill trek from the nearest village. Lalitha Ravi, 52, from Thrissur, had come to attend her grandchild's "choroonu" (rice feeding ceremony) at the Ganapathi temple in Pamba, when rumours over her age triggered protests at the Sannidhabam.
Black flags were shown to Devaswom Minister Kadakampally Surendran at Guruvayur and Health minister K K Shylaja at Kannur by the BJP and its youth wing activists.
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