In a study examining protein denaturation, which factor is most likely to lead to irreversible denaturation of bovine serum albumin in a laboratory setting?

Irreversible denaturation of bovine serum albumin most often occurs when the protein is exposed to extreme heat, such as boiling at 100 °C for several minutes. At these temperatures the disulfide bonds and hydrophobic core are disrupted, causing the protein to unfold and aggregate into insoluble clumps that cannot refold. This loss of native structure is permanent because the unfolded chains form new covalent or non‑covalent contacts that block the re‑formation of the original fold. For example, heating a 1 mg/mL albumin solution to 100 °C for 10 minutes in a water bath produces a visible precipitate that remains insoluble even after cooling. Thus, high temperature is the most likely laboratory factor to cause irreversible denaturation of bovine serum albumin.