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Parah 7:6 describes how the Sanhedrin was ruling on practical matters related to creating mei ḥatat even after the destruction of the second Temple, and it follows that mei ḥatat was still being created and used for purification at this time.

Is this the latest reference of creation/use of mei ḥatat? If not, what is the latest such reference? Is there any reference speaking about the cessation of such use?

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Sefer Hatteruma addresses this issue (closely imitated in Hilchot Erets Yisra'el attributed to the Tur). They bring a few examples that prove that the ashes of the red cow were still in use after the destruction of the Temple, until well into the days of the amora'im, even though it must have been made while the Temple still stood.

  • Nidda 32a: Rabbi Yosi, Rabbi and Rav Yosef all mention cases of immersing a newborn girl who became nidda, so that she could touch teruma or be anointed with teruma.

  • Chullin 107b: Rabbi Ami and Rabbi Asi ate teruma with wineskins so as not to contaminate the teruma with their hands. If there were no ashes of the red cow, they would have been impure in other ways and wouldn't have been concerned only with the impurity of their hands.

  • Nidda 6b: In the days of Rabbi, there were still people who were careful to keep their food pure to be used by the Temple when rebuilt.

  • Bechorot 27b: Rav Nachman asks rhetorically "Do we have the sprinkling of the red cow?" meaning that in Babylon they didn't have it, but in Israel they still had it (Tosafot there seems to have assumed the same).

Interestingly, none of these cases actually says anything about the ashes of the red cow. The Sefer Hatteruma deduces that it must have existed for legal reasons. Otherwise, these aren't exactly direct proofs. Since I assume he would have quoted any explicit statements if there were any rather than relying on the implication of these passages, there are probably no more explicit statements about the amora'im using the ashes of the red cow.

In summary, I think that it's safe to say that many people in the times of the early amora'im they were still careful about purity of their food, and that some rishonim took this as proof that the ashes of the red heifer were still in use at that time.

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    It's important to note that these examples are all in Israel. In Babylon a) the land is all impure by rabbinic decree, and b) the teruma is only rabbinic and could be eaten after mikvah without ashes. – Double AA Sep 19 at 11:27
  • judaism.stackexchange.com/a/55134/759 would probably be another example – Double AA Sep 19 at 11:29
  • Couldn't they just never have become tamei in the first place? Especially the newborn girl. – Heshy Sep 19 at 12:00
  • @Heshy הא על כרחין היו טמאין אי משום אהל המת או משום חרב כחלל של אהל או משום היסח דעת דבעי הזאת שלישי ושביעי כדאיתא פרק טבול יום או משום שבאו מבבל... judge for yourself whether you find it convincing – b a Sep 19 at 13:03
  • @Heshy if that girl was the only eating trumah that would have been pretty notable. Imagine if today some kohein made a point of having a baby in a cave and then smearing freshly hukhshar terumah oil on it. That would be fine, but it wouldn't be normal or expected by any news consumer or manufacturer of oil. Does the Gemara sound like that case was so unusual? Why did they bother keeping tahor terumah oil around for such a bizarre circumstance? – Double AA Sep 19 at 13:14

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