Djelomična karakterizacija frakcije male molekularne mase s krioprotektivnim svojstvom, izolirane iz plašta Humboldtove lignje (Dosidicus gigas)

Zamrzavanje utječe na funkcionalnost proteina ribe uslijed njihove denaturacije i agregacije. Usprkos tome, funkcionalnost strukturnih proteina Humboldtove lignje (Dosidicus gigas) ostaje ista i nakon zamrzavanja, vjerojatno zbog prisutnosti spojeva male molekularne mase s krioprotektivnim svojstvom. Zbog toga smo pomoću Fourier transformirane infracrvene spekroskopije (FTIR) ispitali frakciju male molekularne mase (<1 kDa) koja je topljiva u vodi, izdvojenu iz plašta Humbodtove lignje. Iz dobivenih spektara određeni su maseni udjeli ukupnih ugljikohidrata, slobodnih monosaharida, slobodnih aminokiselina i amonijevog klorida. Svojstvo krioprotekcije i kriostabilnost proteina kojima pridonose spojevi male molekularne mase ispitani su diferencijalnom pretražnom kalorimetrijom. Djelomičnom je karakterizacijom utvrđeno da su glavni sastojci frakcije bili: slobodne aminokiseline (18,84 mg/g), ugljikohidrati (67,1 µg/mg), među kojima i monosaharidi (ukupno 51,1 µg/mg glukoze, fukoze i arabinose), te amonijev klorid (220,4 µg/mg). Najzastupljenije aminokiseline bile su: arginin, sarkozin i taurin. Spojevi male molekularne mase, u masenom udjelu u kojem su prirodno prisutni u plaštu lignje, snizili su ledište vode na -1,2 °C, te povećali temperaturu kristalizacije na 0,66 °C. Stabilnost miofibrilarnih proteina nakon zamrzavanja i odmrzavanja bitno je povećana u prisutnosti spojeva male molekularne mase u usporedbi s kontrolom (proteinima mišića iz kojih su izdvojeni spojevi male molekularne mase), što potvrđuje kriostabilizacijsko svojstvo strukturnih proteina lignje. Osmoliti prisutni u frakciji male molekularne mase inhibirali su denaturaciju i agregaciju proteina te rekristalizaciju leda, čime je održana struktura mišića pri zamrzavanju. Spojevi male molekularne mase pridonijeli su kriostabilnosti mišića čak i pri njihovom malom masenom udjelu.
Freezing conditions affect fish muscle protein functionality due to its denaturation/aggregation. However, jumbo squid (Dosidicus gigas) muscle protein functionality remains stable even after freezing, probably due to the presence of low-molecular-mass compounds (LMMC) as cryoprotectants. Thus, water-soluble LMMC (<1 kDa) fraction obtained from jumbo squid muscle was evaluated by Fourier transform infrared spectrometry. From its spectra, total carbohydrates, free monosaccharides, free amino acids and ammonium chloride were determined. Cryoprotectant capacity and protein cryostability conferred by LMMC were investigated by differential scanning calorimetry. Fraction partial characterization showed that the main components are free amino acids (18.84 mg/g), carbohydrates (67.1 μg/mg) such as monosaccharides (51.1 μg/mg of glucose, fucose and arabinose in total) and ammonium chloride (220.4 μg/mg). Arginine, sarcosine and taurine were the main amino acids in the fraction. LMMC, at the mass fraction present in jumbo squid muscle, lowered the water freezing point to –1.2 °C, inhibiting recrystallization at 0.66 °C. Significant myofibrillar protein stabilization by LMMC was observed after a freeze-thaw cycle compared to control (muscle after extraction of LMMC), proving the effectiveness on jumbo squid protein muscle cryostability. Osmolytes in LMMC fraction inhibited protein denaturation/aggregation and ice recrystallization, maintaining the muscle structure stable under freezing conditions. LMMC conferred protein cryostability even at the very low mass fraction in the muscle.