2.2.2. FTIR

Main Characteristic Peak

All Col-TPU composite nanofiber membranes have five characteristic peaks that were similar to collagen peaks (Figure 3A,B). The amide A band of collagen was near 3315 cm<sup>−</sup>1, the amide B band was at 2920–2944 cm<sup>−</sup>1, the amide I was at 1625−1690 cm<sup>−</sup>1, the amide II band was at 1500−1600 cm<sup>−</sup>1, and the amide III band was oriented at 1200−1300 cm<sup>−</sup>1. Figure 3B and Table S1 shows that the amide A band of Col100 was oriented at 3311 cm<sup>−</sup>1, and the Col-TPU composite nanofiber membranes were blue-shifted from 3307 cm<sup>−</sup><sup>1</sup> to 3309 cm<sup>−</sup><sup>1</sup> with increasing TPU, which may be caused by the N-H vibration (3310 cm<sup>−</sup>1) coupling effect of the group in TPU. The amide B band reflects the ubiquitinated coupling between the amide A band and the amide II band. Col100 was oriented at 2932 cm<sup>−</sup>1, and Col-TPU composite nanofiber membranes were blue-shifted to 2939 cm<sup>−</sup><sup>1</sup> with increasing TPU. This affected the asymmetric stretching of the C-H group (2942 cm<sup>−</sup>1) in TPU, thus indicating that collagen and TPU were prepared successfully. The amide I and II bands of Col100 were oriented at 1655 cm<sup>−</sup><sup>1</sup> and 1655 cm<sup>−</sup>1. The amide I and II bands of Col-TPU composite nanofiber membranes were ultimately red-shifted to 1554 cm<sup>−</sup><sup>1</sup> and 1534 cm<sup>−</sup><sup>1</sup> with increasing TPU ratios. This may be due to the combination of collagen and TPU, thus showing the characteristic peak of TPU and resulting in a red-shift of the hydroxyl peak. The C=C stretching vibration absorption peak (1532 cm<sup>−</sup>1) of TPU in the Col-TPU composite nanofiber membranes approached the N-H out-of-plane vibration absorption peak at 1537 cm<sup>−</sup><sup>1</sup> in collagen, thus leading to two peaks that expanded into one wide absorption peak on the surface of Col-TPU composite nanofiber membranes [25].

The amide III band of Col100 was oriented at 1223 cm<sup>−</sup>1, and the Col-TPU composite nanofiber membranes were blue-shifted to 1230 cm<sup>−</sup><sup>1</sup> upon the addition of TPU, which may affect the vibration of C-C-N in the ethyl carbamate bond (-NHCOO-) in TPU near 1240 cm<sup>−</sup>1. Moreover, the absorption peaks in the region of 1100 cm<sup>−</sup><sup>1</sup> (stretching vibration of C-O-C) in TPU were blue-shifted from 1000 cm<sup>−</sup><sup>1</sup> to 1100 cm<sup>−</sup><sup>1</sup> when the ratio is 60:40 (collagen:TPU). This suggests that TPU combined with collagen to form Col-TPU composite nanofiber membranes during the interfacial interaction between collagen and TPU [26]. The absorption ratio of the amide III band to 1450 cm<sup>−</sup><sup>1</sup> (amide III/A1450) is an important index to determine the integrity of the collagen triple helix structure. When the ratio is <0.5, collagen unwinds the triple helix structure due to denaturation. The absorption ratio of amide III/A1450 in Col100 was 1.114. After adding TPU, and the absorbance ratios of Col-TPU composite nanofiber membranes were 1.131 (Col95), 1.127 (Col90), 1.036 (Col80),

and 0.948 (Col60). These data indicate the presence of triple-helical structures of collagen in Col-TPU composite nanofiber membranes [27].

**Figure 3.** Spectroscopy analysis. (**A**) FTIR of collagen; (**B**) FTIR of Col-TPU composite nanofiber membranes; (**C**) Conformation relative content of amide I band in Col-TPU composite nanofiber membranes; (**D**) XRD of Col-TPU composite nanofiber membranes.
