Droplet and creaming stability of fish oil-loaded gelatin/surfactant-stabilized emulsions depends on both the adsorption ways of emulsifiers and the adjusted pH

Ting Zhang,Mengzhen Ding,Xichang Wang,Jian Zhong

National R&D Branch Center for Freshwater Aquatic Products Processing Technology(Shanghai),Integrated Scientific Research Base on Comprehensive Utilization Technology for By-Products of Aquatic Product Processing,Ministry of Agriculture and Rural Affairs of the People’s Republic of China,Shanghai Engineering Research Center of Aquatic-Product Processing and Preservation,College of Food Science&Technology,Shanghai Ocean University,Shanghai,201306,China

ABSTRACT The droplet and creaming stability of food emulsions stabilized by mixed emulsifiers is a research hotspot in the field of emulsions.In this work, we mainly explore the effect of pH change on the droplet and creaming stability of fish oil-loaded emulsions synergistically(Span 80 and SL)or competitively(Tween 80 and SDS) stabilized by gelatin/surfactant.The results demonstrated that initial droplet stability and droplet storage stability, and creaming stability of the pH-adjusted emulsions are dependent on both the adsorption ways of emulsifiers and the adjusted pH: (1) Competitively stabilized emulsions have more stable droplets than synergistically stabilized emulsions;(2)SDS-dominant competitively stabilized emulsions have more stable droplets than gelatin-dominant emulsions;(3)Basic pH-adjusted emulsions have more stable droplets than acidic pH-adjusted emulsions;(4)The synergistically stabilized emulsions at acidic pH have significantly higher creaming indexes than that at basic pH;and(5)The competitively stabilized emulsions have high or similar creaming indexes to that at acidic pH.Further,the mechanisms are proposed according to Stokes’ Law.This work will provide useful information to understand the interfacial properties of mixed emulsifiers in the food emulsions and promising application perspective for the development of food beverages including acidic and alkali beverages.

Keywords:Emulsion Gelatin pH change Storage Surfactant

1.Introduction

Emulsions are suspensions in which small spherical droplets are dispersed into an immiscible liquid phase[1].They can efficiently encapsulate and deliver biologically active substances such as fish oils for application in food and pharmaceutical fields[2].Emulsions are thermodynamically unstable.Gravitational phase separation,flocculation, coalescence, and Ostwald ripening generally appear during the storage process,which limits their wider application in the food industry[3,4].

The development and application of emulsifiers has attracted much attention to increasing emulsion stability in the past two decades[5,6].Amphiphilic macromolecules can be used to decrease interfacial tension and to form steric elastic films [7].As a typical amphiphilic macromolecule, gelatin has been applied to stabilize emulsions because of its excellent biocompatibility and biodegradability.Gelatin is produced by partial hydrolysis of native collagens[8,9].Both collagens and gelatins have been widely explored in the fields of food [10-12], pharmaceutics [13,14], bioimaging [15,16],and tissue engineering[17-20].Recently,acid or alkali pretreated gelatins [21], crosslinked gelatin particles [22], gelatin/chitosan complexes [23], or gelatin/glucomannan/tannic acid nanocomplexes [24] have been developed to increase the stability of emulsions.

Another type of widely applied emulsifier is low molecular weight surfactant [25].Proteins and surfactants have been investigated for use together to stabilize emulsions.These investigations have found that synergetic interfacial adsorption of protein(caseinate) and surfactants (sodium stearoyl lactylate, phospholipid, and sucrose ester) occurred in aerated emulsions [26] and competitive interfacial adsorption of milk protein and surfactants (sucrose stearate 370, mono- and di-acylglycerols, sucrose stearate 1670, and Tween 20) occurred in frozen aerated emulsions [27].Recent work also demonstrated that four types of surfactants and gelatin were synergistically(Span 80 and soybean lecithin [SL]) and competitively (Tween 80 and sodium dodecyl sulfate [SDS]) adsorbed on the oil/water interfaces in the fish oilloaded gelatin/surfactant-stabilized emulsions [28].Other work also suggested that the droplets and emulsion forms of these emulsions were affected by preparation pH [29].Many studies have demonstrated that the droplet coalescence stability of emulsions stabilized by mixed emulsifiers,especially mixtures of proteins and small molecule surfactants,was worse than that of emulsions stabilized fromeitherindividualemulsifier[4].However,the droplet and creaming stability differences between synergistically stabilized emulsions and competitively stabilized emulsions remain unclear.

The amphiphilic macromolecule and ionic small molecule surfactant emulsifiers may show different electrostatic properties at different pH because they have different isoelectric points [30],which might affect the droplet and creaming stability of the emulsions.Based on the acid-base properties, foods can be classified into alkaline, neutral, and acidic foods.Therefore, the droplet and creaming stability of food emulsions at different adjusted pH should be considered before they are applied for novel alkaline,neutral, and acidic food developments.Therefore, the purpose of this study is to analyze the effect of pH change on the droplet and creaming stability of fish oil-loaded emulsions synergistically(Span 80 and SL) or competitively (Tween 80 and SDS) stabilized by gelatin/surfactants.First, the pH of the freshly prepared gelatin/surfactant-stabilized emulsions will be adjusted to designated values(pH 3,5,7,9,and 11)and the droplet changes will be analyzed.Second,the storage droplet and creaming stability of the pH-adjusted emulsions will be examined by digital camera, optical microscopy, and confocal laser scanning microscopy (CLSM).Finally, the relationship between creaming stability and droplet stability will be discussed according to Stokes’ Law.All results demonstrate that pH change has different effects on the droplet stability of synergistically or competitively stabilized emulsions.

2.Materials and methods

2.1.Materials

Deep sea fish oils (food grade, DHA+EPA ≥70%, Xi’an LvTeng Biological Technology Co., Ltd., Shaanxi Province, China) were stored at-18°C.Bovine bone gelatin granules(type B,240 g bloom,Aladdin Industrial Corp.,Shanghai,China),dyes(Nile Red,Nile Blue,Sangong Biotech.Co., Ltd., Shanghai, China), and other common chemicals(Sinopharm Chemical Reagent Co.Ltd.,Shanghai,China)were stored at room temperature.

2.2.Preparation and pH change of fish oil-loaded gelatin/surfactant-stabilized emulsions

0.8%gelatin solution was incubated in an oscillating(180 r/min),constant-temperature (45°C) water bath (Model SW22, Julabo,Germany).After 60 min, surfactants (Span 80, SL, Tween 80, or SDS) were added into the gelatin solution with a final surfactant concentration of 2.0%.The solution pH was adjusted to pH 11 by using 1 mol/L HCl and 1 mol/L NaOH.Then,fish oils(equal volume)were added into the solutions.The resultant solutions(10 mL)were mechanically sheared by a T 10 basic ULTRA-TURRAX®homogenizer(IKA,Guangzhou,China)with a 10 mm head,a homogenizing speed of 8000 r/min, and a homogenizing time of 120 s.Then, the emulsion pH was adjusted to the designated pH by adding appropriate amounts of HCl or NaOH.The adding amounts were obtained by pre-determined experiments:approximately 80,40,10 μL HCl(1 mol/L), or 20, 40 μL NaOH (1 mol/L) was added into the freshly prepared emulsions to adjust the pH to the designated values(3,5,7,9,or 11).After adding,the emulsions were immediately mechanically sheared with a homogenizing speed of 8000 r/min and a homogenizing time of 10 s to ensure the pH was uniformly changed to the designated pH as quickly as possible.In order to decrease the pH adjustment times and adjustment circles, the volumes of 1 mol/L HCl solution or 1 mol/L NaOH solution were obtained by pre-measuring the added volumes to the same batch of the emulsions.The obtained emulsions were imaged by a digital camera[31].

2.3.Calculation of creaming index

The creaming index (CI) was calculated using the following equation[32]:

Where Hsis the height of the serum layer (the sum of the transparent and/or turbid layers at the bottom of the vials)and Htis the total height of the emulsions.

2.4.Optical microscopy measurements

3 μL liquid emulsion or about 0.003 g emulsion gel samples were put onto glass microslides and were covered by square cover glasses(22 mm×22 mm,extra thin).The samples were observed by an inverted optical microscope(MS600 F,Shanghai Minz Precision Instruments Co.Ltd.,Shanghai,China).

2.5.Gaussian fitting

The droplet sizes in the optical microscopy images were measured by MeizsMcs 6.0 software (Shanghai Minz Precision Instruments Co.Ltd.,Shanghai,China).The frequency distribution of the droplet sizes with a bin size of 1 or 2.5 μm from three batches of the emulsions(about 500-5000 droplets)was statistically analyzed [33].Single or multiple peak Gauss fit was used to analyze the peaks[34].

2.6.Confocal laser scanning microscopy(CLSM)measurements

For liquid emulsions,1 mL emulsion and 40 μL fluorescent dyes(0.1%Nile red and 1%Nile blue)were mixed.They were vortexed for 30 s and were incubated for 5 min in a dark place.To prepare CLSM samples, 3 μL emulsions were transferred onto glass microslides and square cover glasses were used to cover onto the emulsions.For emulsion gels,a 1.0 g gel sample was cut into small pieces with a diameter of about 0.2 mm.The small pieces and 40 μL of the fluorescent dyes (0.1% Nile red and 1% Nile blue) were mixed.They were vortexed for 2 min and incubated for 10 min in a dark place.To prepare CLSM samples, 0.003 g samples were put onto glass microslides and square cover glasses were placed over the small pieces.

These CLSM samples were examined by a confocal laser scanning microscope(TCS SP8,Leica,Wetzlar,Germany)with a 63×oil objective.Nile Red and Nile Blue were excited by 552 and 633 nm laser,respectively[35].The image capture and analysis were performed by Leica Application Suite X software.The operational parameters were as follows:scanning frequency,200 Hz;scanning density,1024×1024.Three parallel samples were analyzed.

Fig.1.Freshly pH-adjusted fish oil-loaded emulsions synergistically(Span 80 and SL)or competitively(Tween 80 and SDS)stabilized by gelatin/surfactant to different pH(3,5,7,9,and 11).(A):Photographs.(B):Optimal microscopy images.All the emulsions are in liquid.Scale bars indicate 50 μm.(C-F):Representative droplet size distribution of the freshly pH-adjusted fish oil-loaded emulsions stabilized by gelatin/Span 80, gelatin/SL, gelatin/Tween 80, and gelatin/SDS, respectively.See Figs.S1-S4 for detailed representative droplet size distribution.

3.Results and discussion

3.1.Initial effect of pH change on the pH-adjusted emulsions

The fish oil-loaded gelatin/surfactant-stabilized emulsions were prepared and the pH was between 7-8 (Data not shown).Then,the emulsion pH was adjusted to the designated pH.None of the emulsions showed obvious creaming separation (Fig.1A).pH change (Fig.1B) had obvious effects on the droplets of the synergistically stabilized emulsions, whereas no obvious effects on the droplets of competitively stabilized emulsions.Further, the droplet sizes of gelatin/Span 80-stabilized, gelatin/SL-stabilized,and gelatin/Tween 80-stabilized emulsions had a typical multimodal distribution,while the droplet sizes of gelatin/SDS-stabilized emulsions had a typical monomodal distribution (Figs.1C-1F, S1-S4).The multimodal and monomodal distributions are consistent with previous work[29].Subsequently,the droplet sizes were compared with those of the freshly prepared emulsions with a gelatin solution pH of 11 in previous work[29].For the synergistically stabilized emulsions (Figs.1C, D, S1 and S2), the acid adding (pH 3,5, and 7) increased the peak 2 and peak 3 droplet sizes, whereas no obvious effects on the peak 1 droplet sizes.In addition, base adding (pH 9 and 11) had no obvious effects on the droplet sizes.It is interesting to note that the acidic pH (pH 3 and 5) increased the droplet size diversity (tetramodal and pentamodal distribution) of gelatin/Span 80-stabilized emulsions.Therefore, droplet coalescences occurred in the synergistically stabilized emulsions.For the competitively stabilized emulsions(Figs.1E,1F,S3 and S4),the acid adding(pH 3,5,and 7)increased the peak 3 droplet sizes of gelatin/Tween 80-stabilized emulsions,whereas pH change had no obvious effects on other droplet sizes.Especially, pH change did not change droplet size diversity.Therefore, the synergistically stabilized emulsions showed stronger pH-dependent droplet coalescence behaviors than the competitively stabilized emulsions.

Fish oil has more than 70%DHA and EPA.The free DHA and EPA are neutrally charged at acidic and neutral pH, and are negatively charged at basic pH.Gelatin’s isoelectric point is at pH 5; it is positively charged at pH 3, neutrally charged at pH 5, and negatively charged at pH 7-11 [29].Span 80 is a type of non-ionic hydrophobic surfactant and is neutrally charged at any pH.SL is an amphiphilic surfactant and is positively charged at acidic pH,neutral at pH 7, and negatively charged at basic pH.Their molecular structures and their synergetic and competitive interactions with gelatins could be referred to our previous work[28,29].In the freshly prepared synergistically stabilized emulsions, gelatin and surfactants(Span 80 and SL)were synergistically adsorbed on the oil/water interfaces.Acid adding to the synergistically stabilized emulsions can change the charges of fish oil,gelatin,and SL.Therefore, acid adding induced the relative electrostatic charges of the shell layer components of the droplets.The different physicochemical stabilization mechanisms against droplet coalescence between gelatin and surfactants (Span 80 and SL) induced droplet coalescences under acidic adjusted pH[36].However,base adding to the synergistically stabilized emulsions does not change the charges of fish oil,gelatin,Span 80,and SL.Therefore,base adding might not induce the droplet coalescences.Previous work has shown that the droplet sizes of gelatin/Span-stabilized, gelatin/SL-stabilized, and gelatin/Tween 80-stabilized emulsions were mainly affected by gelatin, whereas the droplet sizes of gelatin/SDS-stabilized emulsions were mainly affected by SDS[28].Thus,initial droplet stability under pH change was dependent on both the adsorption ways of emulsifiers and the adjusted pH: (1) Competitively stabilized emulsions had more stable droplets than synergistically stabilized emulsions; (2) SDS-dominant competitively stabilized emulsions had more stable droplets than gelatin-dominant emulsions;and(3)Basic pH-adjusted emulsions had more stable droplets than acidic pH-adjusted emulsions.It should be noted that all these droplets were order of micron.Considering that nanometer droplets were more susceptible than larger ones due to Ostwald ripering[37,38],we did not explore the effect of pH on the emulsions with nanometer droplets.

Fig.2.Storage of pH-adjusted fish oil-loaded emulsions synergistically stabilized by gelatin/Span 80 to different pH(3,5,7,9,and 11)at room temperature.Black arrows and asterisk indicate redispersible emulsion gels.Other emulsions are in liquid.Scale bars indicate 50 μm.

3.2.Creaming and droplet stability of the pH-adjusted emulsions

The synergistically stabilized emulsions were changed to emulsion gels during the storage process (Figs.2A and 3A).The gel change time increased with the increase of the adjusted pH.In addition, these emulsions showed different degrees of creaming.For the gelatin/Span 80-stabilized emulsions(Fig.S5),the adjusted pH of 3-5 induced high creaming with final creaming indexes of about 32.2%-37.3%after 21 days storage,whereas the adjusted pH of 7-11 induced low creaming with final creaming indexes of about 7.3%-8.8% after 21 days of storage.For the gelatin/SL-stabilized emulsions (Fig.S6), the adjusted pH of 3-5 induced high creaming with final creaming indexes of about 23.1%-23.9%after 21 days of storage, the adjusted pH of 7 induced moderate creaming with final creaming index of about 18.9% after 21 days of storage, and the adjusted pH of 9-11 showed low creaming with final creaming indexes of about 0.0%-4.4%after 21 days of storage.Therefore,the creaming indexes were mainly dependent on the adjusted pH(acidic pH ＞basic pH) and minorly dependent on the emulsifiers(the creaming at pH 7).These emulsions showed the presence of a yellow layer after creaming(Figs.2A and 3A),which might have resulted from the presence of gelatin molecules.Compared to acid adding(Figs.2B and 3B),base adding(pH 9 and 11)showed a slower transformation from spherical droplet shape to irregular droplet shape to irregular shape,which suggested that droplet coalescence occurred during the storage process.

The competitively stabilized emulsions did not change to emulsion gels even after 21 days of storage (Figs.4A and 5A).These emulsions showed different degrees of creaming(Figs.S7-S8).For the gelatin/Tween 80-stabilized emulsions (Fig.S7), the adjusted pH of 3-5 and 9-11 induced high creaming with final creaming indexes of 34.1%-37.8% after 21 days of storage, whereas the adjusted pH of 7 induced low creaming with final creaming indexes of about 4.3%after 21 days of storage.For the gelatin/SDS-stabilized emulsions(Fig.S8),the adjusted pH of 3-5 induced high creaming with final creaming indexes of about 36.4%-37.7%after 21 days of storage, and the adjusted pH of 7-11 induced moderate creaming with final creaming indexes of about 28.7%-30.8% after 21 days of storage.Therefore,the competitively stabilized emulsions showed no obvious differences in creaming indexes(＞28.7%)with the exception of the gelatin/Tween 80-stabilized emulsion with an adjusted pH of 7.These emulsions showed the presence of a yellow layer at a pH of 3-5 and the presence of a dark brown layer at a pH of 7-11 after creaming (Figs.4A and 5A), which might have resulted from the presence of gelatin molecules.The gelatin/Tween 80-stabilized emulsions at the adjusted pH of 3 (Figs.4B and 5B)had a significant droplet size increase and droplet shape transformation into irregular spherical shapes after 7 days of storage,which was significantly slower than that of the synergistically stabilized emulsions (Figs.2B and 3B).The other competitively stabilized emulsions only showed slight droplet size increases and did not show obvious droplet shape transformation even after 21 days of storage (Figs.4B and 5B).Therefore, slower droplet coalescences occurred in the competitively stabilized emulsions compared with synergistically stabilized emulsions.

Fig.3.Storage of pH-adjusted fish oil-loaded emulsions synergistically stabilized by gelatin/SL to different pH (3, 5, 7, 9, and 11) at room temperature.Black arrows and asterisk indicate redispersible emulsion gels.Other emulsions are in liquid.Scale bars indicate 50 μm.

Because the adjusted pH of 9 induced typical differences for the synergistically and competitively stabilized emulsions after 21 days of storage (Figs.2-5), the emulsions at the adjusted pH of 9 were examined by CLSM(Fig.6).The freshly pH-adjusted emulsions consisted of spherical droplets with different sizes and a fish oil core and a gelatin microstructure shell.After 21 days of storage,the synergistically stabilized emulsions showed irregular shapes,whereas competitively stabilized emulsions only showed slight droplet size increases.These results further confirmed the optical microscopy results (Figs.2-5).According to the merge images, the droplet shell layers consisted of both gelatin and fish oil.Considering small molecule surfactants were also present in the shell layers,the shell layers consisted of both emulsifiers (gelatin and surfactants) and fish oil.

After pH adjustment, the droplet coalescence occurred in the acidic synergistically stabilized emulsions.Without the high energy treatment (homogenization) to form stable droplet microstructures, the size-increased droplets after pH change were not stable and would easily induce droplet coalescences compared to size-unchanged droplets.Therefore, similar to the initial droplet stability under pH change, the storage stability of droplets in the pH-adjusted emulsions was dependent on both the adsorption ways of emulsifiers and the adjusted pH: (1) Competitively stabilized emulsions had more stable droplets than synergistically stabilized emulsions; (2) SDS-dominant competitively stabilized emulsions had more stable droplets than gelatin-dominant emulsions; and (3) Basic pH-adjusted emulsions had more stable droplets than acidic pH-adjusted emulsions.In addition,the creaming stability of the pH-adjusted emulsions was dependent on both the adsorption ways between emulsifiers and the adjusted pH:(1)For the synergistically stabilized emulsions,the creaming indexes of the emulsions at acidic pH were significantly higher than that of the emulsions at basic pH;and(2)For the competitively stabilized emulsions,the creaming indexes of the emulsions at acidic pH were high and were similar to that of the emulsions at basic pH.

Approximation of the move rate of an isolated spherical droplet in an ideal liquid is described by Stokes’Law[4]:

Here,g is the acceleration due to gravity,r is the droplet radius,ρ2is the density of the dispersed phase(droplets in this study),ρ1is the density of the continuous phase (water in this study), and η1is the shear viscosity of the continuous phase(water in this study).The affecting factors for creaming velocity can be described by Eq.(2).

Fig.4.Storage of pH-adjusted fish oil-loaded emulsions competitively stabilized by gelatin/Tween 80 to different pH(3,5,7,9,and 11)at room temperature.All the emulsions are in liquid.Scale bars indicate 50 μm.

The density(ρ2)of an isolated spherical droplet can be described by the following equation[39]:

Here,ris initial droplet radius,ρcoreis the density of the material core inside the droplets (pure fish oil), ρshellis the density of the interfacial layer(stabilizers),and δ is the thickness of the interfacial layer.ρproteinwas taken as 1350 kg/m3[40].ρfishoilwas taken as 930 kg/m3[41].

Droplet aggregation (flocculation or coalescence) can also lead to more rapid creaming [42].The mixed stabilizers may change creaming stability in terms of initial droplet size,density contrast,and inhibition of aggregation[4].

For the synergistically stabilized emulsions, the initial droplet sizes under adjusted and neutral pH were significantly higher than those under basic pH(Fig.1).Moreover,droplet aggregation(coalescence) occurred under adjusted acidic pH (Figs.1-3), which might lead to similar fast creaming.Previous work has shown that the droplet sizes of synergistically stabilized emulsions were mainly affected by gelatin [28].Therefore, the shell layer of these emulsions mainly consist of gelatin and minorly consist of Span 80 or SL.Fish oil is neutral under adjusted neutral pH of 7, which induced that some fish oils left the shell layers(gelatin was still negatively charged),and therefore ρshellincreased.According to Eq.(3),ρ2became close to ρ1.Therefore, the creaming index differences were mainly dependent on the different initial droplet sizes at different adjusted pH(acidic pH ＞basic pH)and minorly dependent on the shell layer density consisted of emulsifiers (increased density at pH 7).

For the competitively stabilized emulsions, the initial droplet sizes under all pH between 3-11 were similar (Fig.1).Moreover, only gelatin/Tween 80-stabilized emulsions at the adjusted pH of 3 had a significant droplet size increase and droplet shape transformation into irregular spherical shapes with time(Fig.4B).Previous work showed that the droplet sizes of gelatin/Tween 80-stabilized emulsions were mainly affected by gelatin,whereas the droplet sizes of gelatin/SDS-stabilized emulsions were mainly affected by SDS [28].Therefore, the shell layer of gelatin/Tween 80-stabilized emulsions mainly consists of gelatin and minorly consist of Tween 80,whereas gelatin/SDS-stabilized emulsions mainly consist of SDS and minorly consist of gelatin.Under an adjusted neutral pH of 7, fish oil changed to neutral, which induced that some fish oil molecules left the shell layers(gelatin and SDS were still negatively charged), and therefore ρshellof these emulsions increased.According to Eq.(3), ρ2became close to ρ1.Therefore,according to Eq.(2),creaming index of gelatin/Tween 80-stabilized emulsions under the adjusted pH of 7 was significantly lower than that of the emulsions under other pHs.The droplet radius of gelatin/SDS-stabilized emulsions was significantly lower than that of gelatin/Tween-stabilized emulsions, and therefore, Vstokesof gelatin/SDS-stabilized emulsions was mainly dependent on the droplet radius, and the creaming index of gelatin/SDS-stabilized emulsions under the adjusted pH of 7 had no obvious change compared to that of gelatin/SDS-stabilized emulsions under other pHs.Therefore,the creaming index showed no obvious differences because of the very similar droplet sizes at different adjusted pHs(CI ＞28.7%) with the exception of the gelatin/Tween 80-stabilized emulsions under the adjusted pH of 7.

Fig.5.Storage of pH-adjusted fish oil-loaded emulsions competitively stabilized by gelatin/SDS to different pH(3,5,7,9,and 11)at room temperature.All the emulsions are in liquid.Scale bars indicate 50 μm.

Fig.6.CLSM images of pH-adjusted fish oil-loaded emulsions synergistically(Span 80 and SL)or competitively(Tween 80 and SDS)stabilized by gelatin/surfactant to pH 9 at 0 h and after 21 days storage.Data scales indicate 25 μm.

4.Conclusions

In this study, the effect of pH change on the droplet stability and creaming stability of fish oil-loaded emulsions synergistically or competitively stabilized by gelatin/surfactant was investigated.The results demonstrated that the initial droplet stability, storage stability of droplets,and creaming stability of the pH-adjusted emulsions were dependent on both the adsorption ways of emulsifiers and the adjusted pH.The initial droplet stability, storage droplet stability, and creaming stability under pH change was dependent on both the adsorption ways of emulsifiers and the adjusted pH: (1) Competitively stabilized emulsions had more stable droplets than synergistically stabilized emulsions; (2) SDSdominant competitively stabilized emulsions had more stable droplets than gelatin-dominant emulsions; (3) Basic pH-adjusted emulsions had more stable droplets than acidic pH-adjusted emulsions.The creaming stability was also dependent on both the adsorption ways between emulsifiers and the adjusted pH: (4)For the synergistically stabilized emulsions,the creaming indexes of the emulsions at acidic pH were significantly higher than that of the emulsions at basic pH; and (5) For the competitively stabilized emulsions, the creaming indexes of the emulsions at acidic pH were high and were similar to that of the emulsions at basic pH.The underlying mechanisms were analyzed according to Stokes’Law.This study will provide useful information for understanding the interfacial properties of food emulsions stabilized by mixed emulsifiers.The development and application of food emulsions stabilized by mixed emulsifiers can also provide potentially promising applications for food and beverages(including acidic and alkali beverages).Further research is necessary to analyze the molecular interfacial properties(zeta potential,surface tension,etc)and their stability of gelatin/surfactant shell layers of the droplets[43,44],the fish oil stability such as oxidative stability,the in vitro and in vivo release of fish oil from the emulsions, and the effect of real food product environments on these emulsions.

Declaration of Competing Interest

The authors declare no conflict of interest.

Acknowledgements

This research has been supported by research grants from the National Key R & D Program of China (No.2019YFD0902003) and Shanghai Municipal Education Commission - Gaoyuan Discipline of Food Science&Technology Grant Support(Shanghai Ocean University).

Appendix A.Supplementary data

Supplementary material related to this article can be found, in the online version, at doi:https://doi.org/10.1016/j.fshw.2020.04.002.