INTRODUCTIONGelatin is a protein substance that contains allessential amino acids except tryptophan. It is formedby cross-links between various polypeptide chainsthat developed after the destruction of the fibrousstructure of collagen pre-treated with acid, alkaline, orenzymes. This protein-based hydrocolloid has a widerange of applications in various industries due to itsunique structural stability, nutritional properties, andother physicochemical characteristics [1]. Particularly,hydrogels and modified gelatin-based compositesare widely used in the food industry, biomedicine,pharmaceuticals, and cosmetology. Gelatin is also usedin the production of food packaging materials due to itsbiocompatibility, biodegradability, non-immunogenicity,and ability to stimulate cell adhesion and proliferation.It can absorb 5–10 times as much water as its weightand is the main ingredient in hard and soft capsules forpharmaceuticals [2–6].There is a high demand for gelatin in the modernmarket of food products and components, as well as inthe pharmaceutical, medical, and cosmetic markets,with an annual average of 326 000 tons producedworldwide. According to Grand View Research, theglobal gelatin market was worth $2.91 billion in 2020.It is estimated to grow by 8% per year and reach about$5 billion by 2025. Russia seeks to produce food gelatindomestically and therefore needs effective technologicaland biotechnological solutions [7–10]. Current researchfocuses on optimizing gelatin production technologiesand searching for new sources of raw materials toreplace the traditional ones (pig skins, bovine skins, andcattle bones).Today, gelatin is still produced with the technologiesdeveloped several decades ago. The process containsthe following stages: pre-treatment of raw materials,extraction of gelatin, processing of gelatin broths,gelatinization, and drying. The efficiency of collagento-gelatin conversion depends on extraction conditions(temperature, time, and pH), concentration, the qualityof raw materials, and their pre-treatment methods.Using chemical solvents for gelatin extraction canresult in a higher gelatin yield along with more lowmolecular-weight protein fragments that will affect thegel’s strength and melting point. However, industrialproduction parameters are not always optimal, leadingto a low gelatin yield. Therefore, we need to search foralternative solutions to optimize the process.Drying is an important process to obtain gelatinwith improved functional properties. These propertiesbasically depend on the type of raw materials, pretreatmentmethods, drying and extraction conditions,as well as the spatial structure of protein molecules andtheir state. Drying causes physicochemical changes inthe structure and functions of proteins. For example,heating, which is part of the drying process, can breakcovalent and non-covalent bonds leading to changes inthe protein structure. If significant, these changes cangreatly affect gelatin’s functional properties such assolubility, gelation, foaming, emulsification, as well asfat and water absorption. The extent of these changesis mainly determined by the drying methods andconditions [11–15].Drying methods used in the production of proteiningredients (including gelatin) are convection drying,infrared drying, spray drying, and freeze drying.Convection is the most common method of fooddrying. In convection drying, a stream of heated air isdirected at a wet sample. The air here is both a heatingagent and a dehydrator, since it carries away moisturevapor from the dryer. As a result of this lengthy processand elevated temperatures, the final product loses asignificant amount of micronutrients and bioactivecompounds. Although this method is simple to use,convection dryers have low productivity which can leadto uneven drying [16, 17].Infrared heating with microwaves is a new methodof heat treatment (drying) that extends shelf life,reduces drying time, and preserves food quality. Themicrowaves transfer water to the surface where itquickly evaporates under the influence of infraredradiation, which reduces the drying time [18, 19].Spray drying is widely used in the food industrydue to its simplicity and short drying time. This methodallows for a good quality powdered product. However,spray drying causes particles to greatly shrink andbecome denser [20, 21].Freeze drying is a process of removing water from aproduct by freezing it and then converting ice into steam.This process consists of three main stages: freezing,primary drying, and secondary drying. Freezing createsa solid matrix suitable for drying. Primary dryingremoves ice by sublimation, when the pressure in thesystem is reduced but the temperature remains the same.Secondary drying removes bound water reducing it toresidual moisture.Several studies indicate that protein denaturationduring the formation of ice crystals can significantlychange the protein structure. Therefore, whenoptimizing the freezing process, we should take intoaccount the ice surface area, since it can contribute toprotein denaturation caused by freezing [22–24].In spray drying, evaporated material is sprayedthrough the nozzles of a conical-cylindrical apparatus(spray dryer) to obtain a product in the form of apowder or granules. This method is used to drysolutions or suspensions. Spray-dried products includepowdered milk, food and fodder yeast, and egg powder.According to some studies, spray drying can effectivelyeliminate many of the shortcomings of protein andbioactive peptides, such as low bioavailability, highhygroscopicity, physical and chemical instability, aswell as strong bitterness during and after storage [13].It is also claimed that this method can improve gelatin’sfunctional properties, compared to freeze or vacuumdrying [25]. Assumingly, various drying methodsaffect the solubility and amphiphilicity of gelatin as ahigh-protein product, thereby changing its functionalproperties [26].We studied the effect of spray and freeze drying onthe solubility and amphiphilicity (hydrophobicity andhydrophilicity) of gelatin which we obtained in theprevious study by enzymatic acid hydrolysis [7].STUDY OBJECTS AND METHODSThe control sample was a commercially producededible gelatin. The experimental samples were sprayandfreeze-dried gelatins obtained by enzymatic-acidhydrolysis of cattle bone. For this, 3 kg of defattedbeef bones was crushed to particles of 3.0 ± 0.5 mm ina laboratory chain grinder. The bones were obtainedfrom a farm in Kuzbass (Russia). The crushed boneswere placed in a solution of hydrochloric acid (1M HCl)which contained pepsin with an enzymatic activityof 300 000 units. The hydrolysis was carried out at27 ± 2°C for 60 to 240 min, with a pH of 1.5–2.0. MS-01magnetic stirrers (ELMI) were used throughout the254Voroshilin R.A. et al. Foods and Raw Materials. 2022;10(2):252–261experiment to stir the bone material at 100 rpm and27 ± 2°C to ensure its uniform treatment with thesolution. The hydrolyzed material was centrifuged ina high-speed Avanti J-26S centrifuge (Beckman) toseparate the mineral sediment from ossein. Then, theresulting ossein was washed with demineralized waterand subjected to gelatin extraction. A detailed schemeof hydrolysis and gelatin extraction is described in ourprevious work [7].Next, the gelatin broths were dried by the spray- andfreeze-drying methods. The spray-dried gelatin wasobtained in a B-290 Mini Spray Dryer (Buchi, Sweden)at 95°С and а rate of 3.0–3.2 mL/min. The freeze-driedgelatin was obtained in an INEY-6M freeze-drier. Forthis, gelatin broth was poured onto pallets in 1-cm layersand placed in the drying chambers. The chambers wereclosed with lids and the refrigerator was turned on. Theunit entered the freezing mode within 15 min and whenthe evaporator temperature reached minus –35°С, thevacuum pump turned on to start the drying mode. Thefreeze-dried gelatin was then ground in an NS-2000automatic laboratory mill.The amino acid sequence of gelatin proteins, whichis represented by a single letter code, was determinedby matrix-activated laser desorption/ionization ona MALDI Biotyper (Bruker). Amino acid residues,isoelectric point, aliphatic index, molar absorptioncoefficient, as well as the surface area of the peptides,were determined by the in silico bioinformatic methodson the PepDraw online server. The gelatin samples’solubility was evaluated visually. For this, 500 mgof gelatin was mixed with 50 mL of distilled waterand stirred actively (200 rpm) with MS-01 magneticstirrers (ELMI). Dissolution was monitored at watertemperatures of 25 and 50°C.The Bloom strength of gelatin gels was determinedon a ST-2 Strukturometr texture analyzer with a Bloomindenter. For this, 7.5 g of gelatin was placed in a glassof cold water (105 mL) and kept at 22°C max for 180min. Next, the swollen gelatin was heated to 60°Cin a water bath and stirred for 15 min until completedissolution. The solution (6.67% concentration) waspoured into a calibrated beaker and kept at 10.0 ± 0.1°Cfor 17 h. The prepared samples were then placed on theanalyzer’s table under the Bloom indenter for the study.The arithmetic mean of two determinations was taken asthe final result.The ProtScale online service was used to predictthe topology of the hydrophobicity and hydrophilicityof gelatin proteins. In particular, this service allowsus to compute and represent (in the form of a twodimensionalgraph) the profile produced by any aminoacid scale for a selected protein. The amino acid scaleis defined by a numerical value assigned to each type ofamino acid. ProtScale uses the Kyte and Doolittle scalethat assigns individual values to 20 amino acids, namelyAla: 1.800, Arg: –4.500, Asn: –3.500, Asp: –3.500, Cys:2.500, Gln: –3.500, Glu: –3.500, Gly: –0.400, His:–3.200, Ile: 4.500, Leu: 3.800, Lys: –3.900, Met: 1.900,Phe: 2.800, Pro: –1.600, Ser: –0.800, Thr: –0.700, Trp:–0.900, Tyr: –1.300, and Val: 4.200, –3.500, –3.500,–0.490.The molecular weight distribution of proteins wascarried out by polyacrylamide gel electrophoresis inthe presence of an anionic detergent, sodium dodecylsulfate (SDS-Na). For this, the dried gelatin sampleswere dissolved in deionized water at 60°C to createa 0.2% solution. The solution was then mixed with aloading buffer containing 5 μL of dithiothreitol (DTT)and subjected to heat denaturation in boiling waterfor 5 min. After that, 15-μL samples were loaded intopolyacrylamide gels containing 6% of separating gel and5% of stacking gel to perform electrophoresis. Then, thegels were stained with 0.1% Coomassie Blue R-250 in25% isopropanol and 10% acetic acid for 2 h, followedby decoloring with 5% alcohol and 10% acetic acid.Next, 2-D gels were detected using the Gel Doc XR PlusBio-RAD system.RESULTS AND DISCUSSIONFirst, we spray- and freeze-dried the experimentalsamples of gelatin obtained by enzymatic acidhydrolysis. Next, we determined the amino acidsequence of all the dried gelatins (Table 1).The proteins of the control, spray-dried, and freezedriedsamples are represented by peptide sequences of85, 93, and 95 amino acids, respectively (Table 1).These sequences allowed us to determine the aminoacid composition (% or g/100 g of total amino acids) ofthe control and experimental samples. This is a criticalindicator of gelatin quality largely depending on rawTable 1 Amino acid sequences of gelatin samples (one-letter coding)*Control Experimental samplesSpray-dried gelatin Freeze-dried gelatinGGPAAGGPAYGGPILILAPAILAPYILAAILADNPAANPAYNPILPNAAPNAYPNILPQGAPQGYSEAASEAYSEILTNAATNAYPATNSHILEILDVILDHILILDMILSHESHPYCGDDGGYGPYPDDPGYDDGYHEHPILMEMPPYQCCGQNYYNCDDENNPQQRRSVYAEVPYQCCVPGGPILEVILEILESHILEMILHILMILSHPSHPEEPEEEMPEMPPRPPRVRPVREPEHPHPILMPMPRPREYPYESGQSQYNADEGNNPPPQQRS*A – alanine; C – cysteine; D – aspartic acid; E – glutamic acid; F – phenylalanine; G – glycine; H – histidine; I – isoleucine; K – lysine;L – leucine; M – methionine; N – asparagine; P – proline; Q – glutamine; R – arginine; S – serine; T – threonine; V – valine; W – tryptophan;Y – tyrosine255Voroshilin R.A. et al. Foods and Raw Materials. 2022;10(2):252–261materials. Glycine and proline are the most importantamino acids in gelatin. Collagen consists of threeidentical or different polypeptide chains with a repeatingpattern (Gly-XY)n (X and Y stand for any amino acid)and a high content of imino acids with a triple helicalstructure due to hydrogen bonds [27–29].The composition and content of amino acids,especially imino acids, in gelatin have a significantimpact on its structure and functional properties. Inparticular, the gel’s supercoil structure is stabilizedby both the hydrogen bonds forming between aminoacid residues and the pyrrolidine rings of imino acids.A higher content of imino acids ensures a higher gelmodulus, gelling temperature, and melting point [30].The amino acid composition (% or g/100 g of totalamino acids) of the control and experimental gelatinsamples are presented in Table 2.We found that the samples varied mostly in thecontent of alanine, accounting for 25.840% in thecontrol sample and only 1% in the experimental samples.None of the samples contained phenylalanine, lysine,or tryptophan. According to literature, the absence oftryptophan is what makes gelatin different from otherhydrocolloids of animal origin. This amino acid ismainly present in membrane proteins and has aromaticresidues in its structure.Histidine, arginine, and threonine were not detectedin the control sample.Using the PepDraw online server, we determinedthe mass of amino acid residues defined as a sum ofmonoisotopic masses of all amino acid residues inthe peptide. We also calculated the isoelectric pointrepresented by a pH value at which the total chargeof the peptide equals zero. This calculation showsthe partial charge of the peptide at various pH values,starting from 0. Then, we determined the aliphatic indexof the protein defined as a relative volume of aliphaticside chains (alanine, valine, isoleucine, and leucine). Itcan be considered a positive factor in increasing thermalstability of globular proteins.The mass of amino acid residues in the control,spray-dried, and freeze-dried samples amounted to13173.86, 10830.72, and 11156.79, respectively.The aliphatic index values in the control, spray-dried,and freeze-dried samples were 95.96, 87.53, and 70.74,respectively.The isoelectric points in the control, spray-dried,and freeze-dried samples were 5.97, 4.89, and 5.96,respectively.The molar absorption coefficients in the control,spray-dried, and freeze-dried samples were 8960.00,12800.00, and 3840.00 M–1∙cm–1, respectively.The surface area values in the control, spray-dried,and freeze-dried samples were 21223.00, 23040.00, and18407.00, respectively.We concluded that the control and the spray-driedsamples had more thermostable proteins, since theiraliphatic indexes (87 and 96, respectively) were higherthan those of the freeze-dried sample (70). The samples’isoelectric points indicated a slightly acidic reaction,therefore their protein molecules were neutral at a pHvalue of 4.89 to 5.97.Solubility is an important property of gelatin in foodsystems. In cold water, gelatin hydrates and swells, andTable 2 Amino acid contents in the control and experimental gelatin samplesAmino acid Content of total amino acids, % or g/100 gControl Spray-dried gelatin Freeze-dried gelatinA – alanine 25.840 1.050 1.080C – cysteine ND 6.320 NDD – aspartic acid 1.120 11.580 1.080E – glutamic acid 3.370 6.320 16.130F – phenylalanine ND ND NDG – glycine 8.990 9.470 2.150H – histidine ND 6.320 6.450I – isoleucine 8.990 7.370 8.600K – lysine ND ND NDL – leucine 8.990 7.370 8.600M – methionine ND 3.160 6.450N – asparagine 10.110 4.210 3.230P – proline 15.730 10.530 21.510Q – glutamine 2.250 5.260 4.300R – arginine ND 2.110 7.530S – serine 3.370 4.210 6.450T – threonine 3.37 ND NDV – valine ND 4.21 3.23W – tryptophan ND ND NDY – tyrosine 7.87 10.53 3.23ND – not detected256Voroshilin R.A. et al. Foods and Raw Materials. 2022;10(2):252–261at temperatures above 40°C, it forms a colloidal solution(sol). The solubility index depends on the methodof gelatin production. New methods are currentlybeing developed to obtain water-soluble gelatin attemperatures below 40°C. Such gelatin usually has anamorphous powdery form.Next, we visually assessed the degree of solubilityof the gelatin samples at water temperatures of 25 and50°C (Fig. 1).As we can see, the control and the spray-driedsamples showed higher protein solubility at 25 and 50°Cthan the freeze-dried sample. According to Fig. 1c andf, gelatin particles did not dissolve after 1 min of mixingat different temperatures, settling on the bottom and onthe surface. After 3 min of mixing at 25 or 50°C, thefreeze-dried sample still did not dissolve completely,its particles settling on the water surface (Fig. 1f). Thiscould be due to the sample’s mechanical grinding in alaboratory mill at the final stage of freeze-drying, whichresulted in larger particles than those in the spray-driedgelatin and affected its solubility. We can also assumethat spray drying exposes protein molecules to lessthermal stress than freeze drying, which causes thehighest degree of thermal and dehydration stress.Next, we evaluated the Bloom strength of the gelatingels (Fig. 2).The Bloom value is an important parameter ofgelatin’s physical and mechanical properties used infood production. It is also used as a criterion in gelatinclassifications.The gel strength index depends on the proteincontent and the molecular weight of peptides formedin gelatin. In our study, this index was quite high inthe control and spray-dried samples, amounting to229.0 ± 0.5 and 224.0 ± 0.5 Bloom, respectively. Thefreeze-dried sample’s index (186.0 ± 0.5 Bloom) was by17 and 19% lower than for the control and spray-driedsamples. Assumingly, the proteins of the freeze-driedgelatin had a lower molecular weight, which worsenedits structural and mechanical properties. We can alsoassume that this sample might have more low-molecularweight (below 20 kDa) peptides.Next, we determined the degree of proteinhydrophilicity and hydrophobicity based on the aminoacid sequences. Using the ProtScale online service (theKyte and Doolittle scale), we predicted the topology ofprotein hydrophobicity and hydrophilicity for the controland experimental gelatin samples (Fig. 3).On the Kyte and Doolittle scale, the peaks above0 refer to hydrophobicity and those below 0 refer tohydrophilicity. As we can see in Fig. 3a, the controlsample had higher hydrophilic properties since its peaksalong the X axis ranged from 3 to 21, with a peptidesequence of PAAGGPAY GGPILILAPA I. Most of itspeaks were for alanine, proline, isoleucine, and glycine.These amino acids had hydrophobic properties and 1 to4 uncharged side radicals at pH = 6–7. In general, thesepeaks characterized a sequence of amino acids withhydrophobic properties.Figure 1 Dissolution of gelatin samples at 25 and 50°C for 1–3 mina b cFreeze-dried sample, t = 25°С1 мин. 3 мин.1 мин. 3 мин.1 мин. 3 мин.a. Контрольный образец, t=25°С b. Опытный образец 1, t=25°С c. Опытный образец 2, t=25°С1 мин. 3 мин.1 мин. 3 мин.1 мин. 3 мин.d. Контрольный образец, t=50°С e. Опытный образец 1, t=50°С f. Опытный образец 2, t=50°С1 мин. 3 мин.1 мин. 3 мин.1 мин. 3 мин.a. Контрольный образец, t=25°С b. Опытный образец 1, t=25°С c. Опытный образец 2, t=25°С1 мин. 3 мин.1 мин. 3 мин.1 мин. мин.d. Контрольный образец, t=50°С e. Опытный образец 1, t=50°С f. Опытный образец 2, t=50°СControl, t = 25°С Spray-dried sample, t = 25°С1 min 3 min 1 min 3 min 1 min 3 minControl, t = 50°С Spray-dried sample, t = 50°С Freeze-dried sample, t = 50°С1 min 3 min 1 min 3 min 1 min 3 mind e f257Voroshilin R.A. et al. Foods and Raw Materials. 2022;10(2):252–261The region from 22 to 53 had a sequence ofLAPYILAAI LADNPAANPA YNPILPNAAP NAY,whereas the region from 56 to 85 was represented byPQGAPQGYSEAAS EAYSEILTNA ATNAY. Thus,21 out of 85 amino acids had hydrophobic properties.According to Fig. 3a (control), leucine had maximumhydrophobicity of 2.585 units at point 17 and asparginhad highest hydrophilicity of –2.678 at 89.Figure 2 Bloom strength of gelatin gels229 224186-2%-17%050100150200250Control Spray-dried sample Freeze-dried sampleGel strength, BloomThe profile of the spray-dried sample is shown inFig. 3b. As we can see, the peptide region from 5 to 21represented by AGGPAY GGPILILAPA I and the regionfrom 55 to 57 with a NIL sequence had hydrophobicproperties. Most of the peaks were located above0 and were represented by alanine, isoleucine, andproline. These amino acids had hydrophobic propertiesand 1 to 3 uncharged side radicals. Leucine (18) had amaximum hydrophobicity value of 2.500 units, while10 20 30 40 50 60 70 80 90Score43110-1-2-3PositioncFigure 3 Predicted topology of protein hydrophobicity and hydrophilicity in the control and experimental gelatins (a – control;b – spray-dried sample; c – freeze-dried sample)ScorePosition10 20 30 40 50 60 70 803210-1-2-3-4a bPosition20 40 60 80 1003210-1-2-3Score258Voroshilin R.A. et al. Foods and Raw Materials. 2022;10(2):252–261glutamine (77) had a maximum hydrophilicity value of –3.511. Thus, 20 out of 91 amino acids had hydrophobicproperties.The profile of the freeze-dried sample is shownin Fig. 3c. As can be seen, the peptide region from5 to 26 was represented by a sequence of VILEILESHILEMILH ILMILS, the region from 32 to 39 hada sequence of EEPEEEMP, and the one from 57 to 61was represented by PHPI. Glutamic acid, isoleucine,and leucine had most peaks in the hydrophobicity area.They also had from 4 to 7 uncharged side radicals atpH = 6–7. Isoleucine (6) had maximum hydrophobicityof 3.856 units, while glutamine (76) had maximumhydrophilicity of –5.504 units. Thus, 36 out of 93amino acids had hydrophobic properties. Our resultswere consistent with literature on the amphiphilic(hydrophobic and hydrophilic) properties of these aminoacids [31–38].The hydrophobicity values based on the amino acidsequences of the gelatin samples confirmed our dataon their solubility. In particular, they proved that themethod of drying affects the gelatin’s structural andmechanical properties, as well as its physicochemicalparameters. Spray drying can improve the proteins’functional properties compared to freeze drying.Therefore, we can conclude that different dryingmethods affect the solubility and amphiphilicityproperties of gelatin, thereby changing its functionalproperties.Finally, we analyzed the molecular weightdistribution of proteins by polyacrylamide gel electrophoresisin the presence of an anionic detergent,sodium dodecyl sulfate (Fig. 4, Table 3).According to the results, the control sample’s proteinfractions were more evenly distributed by molecularweight compared to the experimental samples. Itsfractions between 50 and 100 kDa accounted for72.6% and those below 20 kDa amounted to 6.1138%of the total content. The spray-dried sample showed asomewhat different molecular weight distribution. Itsprotein fractions between 40 to 100 kDa made up 73.8%,while those below 20 kDa accounted for 5.026315%.The freeze-dried sample had a completely differentdistribution of protein fractions. Most peptides werefound at the level of 40 kDa (42.83855%). Yet, thissample had 30.214499% of proteins with a molecularweight below 20 kDa, which was by 20.23% more thanin the control and by 16.64% more than in the spraydriedgelatin. This increase in low-molecular weightpeptides by an average of 18% was most likely causedby the cleavage of peptides during freeze drying.Our results showed that the degree of degradationof gelatin components can depend on the methodof drying gelatin broths. Freeze drying can lead tomaximum degradation, which may be associated withlong heat treatment (5 h). This time is much longercompared to spray drying, although the process offreeze drying takes place at a lower temperature (60°C).Also, temperatures below 0°C cause gelation followedby freezing, which can also lead to structural changesin gelatin. In addition, a faster process of spray dryingcan slow down the degradation of gelatin proteins.These results were consistent with those for gelatinsolubility (Fig. 2). Therefore, spray drying is moresuitable for maintaining the structure of gelatin and itsfunctional properties.CONCLUSIONWe studied the effect of spray and freeze dryingof gelatin broths on the solubility and amphiphilicityof gelatin. The results showed that spray drying canreduce the breakdown of gelatin proteins and retainmore α-chains, while freeze drying increases thehydrophobicity of gelatin and decreases its solubility.The predicted topology of protein hydrophobicity, whichwas based on the amino acid sequences of the gelatinsamples, confirmed the results on solubility. Particularly,the freeze-dried gelatin had 36 amino acids withhydrophobic properties out of 93, compared to 21 outof 85 in the control and 20 out of 91 in the spray-driedsample.We found that the freeze-dried sample had by 18%more low-molecular weight peptides (below 20 kDa)compared to the control and the spray-dried samples.This was most likely caused by the cleavage of peptidesduring the drying process. Freeze drying can lead toFigure 4 Electropherogram of the molecular weightdistribution of gelatin samples (1 – marker; 2 – control,3 – spray-dried sample, 4 – freeze-dried sample)200 кДа150 кДа100 кДа85 кДа60 кДа50 кДа40 кДа30 кДа25 кДа20 кДа1 2 3 4kDa20253040506085100150200259Voroshilin R.A. et al. Foods and Raw Materials. 2022;10(2):252–261maximum degradation of gelatin components due tolong heat treatment. Temperatures below 0°C causegelation followed by freezing, which can also causestructural changes in gelatin. By contrast, a faster spraydrying process can, to a certain extent, slow down thedegradation of gelatin proteins.Thus, spray drying is more suitable for gelatindrying, since this method improves the stability ofgelatin’s outer and inner structure, which was confirmedby high hydrophilicity values of the spray-dried sample.Table 3 Molecular weight distribution of gelatin samplesMolecular weight, kDa Molecular weight distribution, %Control Spray-dried sample Freeze-dried sample200 1.316515 3.878759 6.386503150 3.101737 5.118362 1.334564100 19.4574 17.810301 1.30143685 17.85029 13.035829 1.36769160 14.53681 8.429303 42.8385550 20.790700 21.727447 3.17077240 10.111660 12.89394 3.45945430 0.220568 6.732710 3.53044225 2.909430 1.157420 3.45235620 3.591122 4.189303 2.974374Below 20 6.113800 5.026315 30.214499Further research could search for optimal parametersand modes of spray drying for gelatin broths.CONTRIBUTIONThe authors were equally involved in writing themanuscript and are equally responsible for plagiarism.CONFLICT OF INTERESTThe authors declare that there is no conflict ofinterest regarding the publication of this article.