Innovation in a Continuous System of Distillation by Steam to Obtain Essential Oil from Persian Lime Juice (Citrus Latifolia Tanaka)

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Article Innovation in a Continuous System of Distillation by Steam to Obtain Essential Oil from Persian Lime Juice (Citrus latifolia Tanaka)

José Daniel Padilla-de la Rosa 1 , Magaly Dyanira Manzano-Alfaro 2, Jaime Rosalío Gómez-Huerta 3, Enrique Arriola-Guevara 2, Guadalupe Guatemala-Morales 4,* , Anaberta Cardador-Martínez 5,* and Mirna Estarrón-Espinosa 1,*

1 Unidad Zapopan, Centro de Investigación y Asistencia en Tecnología y Diseño del Estado de Jalisco (CIATEJ), Av. Camino Arenero No. 1227, Col. El Bajío, Guadalajara C.P. 45019, Mexico; [email protected] 2 Departamento de Ingeniería Química, CUCEI, Universidad de Guadalajara, Blvd. Marcelino García Barragán #1421, esq. Calzada Olímpica, Guadalajara C.P. 44430, Mexico; [email protected] (M.D.M.-A.); [email protected] (E.A.-G.) 3 Departamento de Química, CUCEI, Universidad de Guadalajara, Blvd. Marcelino García Barragán #1421, esq. Calzada Olímpica, Guadalajara C.P. 44430, Mexico; [email protected] 4 Unidad Normalistas, Centro de Investigación y Asistencia en Tecnología y Diseño del Estado de Jalisco (CIATEJ), Av. Normalistas No. 800, Guadalajara C.P. 44720, Mexico 5 Escuela de Ingeniería y Ciencias, Tecnologico de Monterrey, Epigmenio González 500 Fracc. San Pablo, Querétaro C.P. 76130, Mexico Citation: Padilla-de la Rosa, J.D.; * Correspondence: [email protected] (G.G.-M.); [email protected] (A.C.-M.); Manzano-Alfaro, M.D.; [email protected] (M.E.-E.) Gómez-Huerta, J.R.; Arriola-Guevara, E.; Guatemala-Morales, G.; Abstract: The citrus industry is one of the most important economic areas within the global agricul- Cardador-Martínez, A.; tural sector. Persian lime is commonly used to produce lime juice and essential oil, which are usually Estarrón-Espinosa, M. Innovation in a obtained by batch distillation. The aim of this work was to validate a patented continuous steam dis- Continuous System of Distillation by tillation process and to both physically and chemically characterize the volatile fractions of essential Steam to Obtain Essential Oil from Persian lime oil. Prior to distillation, lime juice was obtained by pressing the lime fruit. Afterwards, Persian Lime Juice (Citrus latifolia the juice was subjected to a continuous steam distillation process by varying the ratio of distillate Tanaka). Molecules 2021, 26, 4172. flow to feed flow (0.2, 0.4, and 0.6). The distillate oil fractions were characterized by measuring their https://doi.org/10.3390/ density, optical rotation, and refractive index. Gas chromatography GC-FID was used to analyze the molecules26144172 chemical compositions of the oil fractions. The process of continuous steam distillation presented high oil recovery efficiencies (up to 90%) and lower steam consumption compared to traditional Academic Editor: Laura De Martino batch process distillation since steam consumption ranged from 32 to 60% for different steam levels.

Received: 11 June 2021 Moreover, a reduction in process time was observed (from 8 to 4 h). The oil fractions obtained via Accepted: 3 July 2021 continuous steam distillation differed signiﬁcantly in their composition from the parent compounds Published: 9 July 2021 and the fractions.

Publisher’s Note: MDPI stays neutral Keywords: essential oils; distillation by steam; continuous processes with regard to jurisdictional claims in published maps and institutional afﬁl- iations. 1. Introduction Citrus is the most abundant crop of fruit trees around the world, with annual production in 2015 totaling approximately 130,947 million tons. The main citrus fruits grown Copyright: © 2021 by the authors. globally are oranges, mandarins, lemons, limes, and grapefruits. Brazil, China, India, Licensee MDPI, Basel, Switzerland. Mexico, Spain, and the USA produce over two-thirds of the world’s citrus fruits [1]. In This article is an open access article 2017, the worldwide production of acid lime fruit was 17,218,173 tons, among which Mex- distributed under the terms and ico contributed 2,528,174 tons and was thus considered the main producer of lime fruit, conditions of the Creative Commons comprising 14.7% of the total production [2]. Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

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1.1. Industrial Use of Lemons

1.1. IndustrialFor many Use years, of Lemons the citrus processing industry has focused on the production of juices and essential oils. It was estimated that 33% of citrus harvested in the world is used For many years, the citrus processing industry has focused on the production of juices to produce juices [3]. In the processing of citrus fruits (orange, lemon, lime, grapefruit, and essential oils. It was estimated that 33% of citrus harvested in the world is used to etc.) for juice and essential oil extractions, peels represent between 50 and 65% of the total produce juices [3]. In the processing of citrus fruits (orange, lemon, lime, grapefruit, etc.) weight of the fruits and remain the primary byproduct [4]. This solid residue, referred to for juice and essential oil extractions, peels represent between 50 and 65% of the total as citrus waste, is estimated to total 15 million tons per year worldwide [5]. Although weight of the fruits and remain the primary byproduct [4]. This solid residue, referred to as citrus peels are used as cattle feed and as a source of essential oils, the correct management citrus waste, is estimated to total 15 million tons per year worldwide [5]. Although citrus of this residue remains an ongoing problem for the citrus industry [3]. The major compo- peels are used as cattle feed and as a source of essential oils, the correct management of nents of dry citrus peel waste include sugars (23%), cellulose (22%), pectin (25%), hemi- this residue remains an ongoing problem for the citrus industry [3]. The major components cellulose (11%), flavonoids (4.5%), and up to 4% essential oil, often referred to as citrus of dry citrus peel waste include sugars (23%), cellulose (22%), pectin (25%), hemicellulose essential oil [6]. (11%), flavonoids (4.5%), and up to 4% essential oil, often referred to as citrus essential oil [6]. 1.2. Production of Essential Oils 1.2. ProductionDifferent industries of Essential appreciate Oils citrus essential oils because of their pleasant aroma and incredibleDifferent industries variety of appreciate applications citrus [7]. essential In the food oils industry, because of citrus their pleasantessential aromaoil is used and asincredible the main variety aromatic of applications ingredient in [7 ].non-alcoholic In the food industry, carbonated citrus drinks, essential cola oildrinks, is used pastries, as the sweets,main aromatic and biscuits. ingredient In the in pharmaceutical non-alcoholic carbonated industry, this drinks, type cola of essential drinks, pastries, oil is used sweets, as a flavoringand biscuits. to mask In the the pharmaceutical unpleasant taste industry, of drugs this [8]. type In ofthe essential chemical oil industry, is usedas these a flavoring oils are usedto mask as fragrances the unpleasant in air tastefresheners of drugs and [8 cleani]. In theng products. chemical industry,In the perfume these oilsindustry, are used citrus as essentialfragrances oils in airare freshenersused as the and primary cleaning raw products. materials In to the formulate perfume industry,toiletries citrusand perfumes essential [7].oils are used as the primary raw materials to formulate toiletries and perfumes [7]. Citrus oil generally contains over 90% terpenes, about 5% oxygenated compounds, and less than than 1% 1% non-volatile non-volatile compounds, compounds, such such as as waxes waxes and and pigments pigments [9,10]. [9,10 ].Essential Essen- oilstial consist oils consist of mixtures of mixtures of volatile of volatile and non-volatile and non-volatile components, components, such as suchterpenes, as terpenes, sesquiterpenes,sesquiterpenes, higher higheralcohols, alcohols, aldehydes, aldehydes, ketones, ketones, acids, esters, acids, esters,camphor, camphor, and waxes. and waxes. The volatilevolatile fractionfraction of of Persian Persian lime lime oil oil is characterizedis characterized by higherby higher levels levels of d-limonene, of d-limonene,γ-terpinene, γ-terpinene, esters, esters, and monoterpene and monoterpene aldehydes aldehydes and lower and amounts lower amounts of β-pinene of β +-pinene sabinene + sabinenesesquiterpenes sesquiterpenes and aliphatic and aliphatic aldehydes aldehydes than key than lime key oils lime [11 oils]. Interestingly, [11]. Interestingly, although alt- houghd-limonene d-limonene comprises comprises more than more 90% thanw/w 90%of the w/w volatile of the compositionvolatile composition of the essential of the essen- oil, its tialcontribution oil, its contribution to the final aromato the final of the aroma mixture of isthe limited mixture by itsis limited instability by andits instability the possibility and theof forming possibility off-flavor of forming compounds off-flavor [7]. compounds [7]. Some ofof thethe representative representative compounds compounds in thein essentialthe essential oil from oil Persianfrom Persian lime are lime shown are shownin Figure in1 Figure. 1.

Figure 1. Compounds in in the the essential essential oil oil of of Persian Persian lime: lime: (a ()a d-limonene,) d-limonene, (b ()b γ)-terpinene,γ-terpinene, (c) (βc) -pinene,β -pinene, (d) (αd-terpineol,) α-terpineol, (e) (neral,e) neral, and and (f) (geranial.f) geranial.

1.3. Types of Essential Oils There are different types of essentialessential oils dependingdepending on the processprocess usedused toto obtainobtain them, including cold-pressed oil, distilled oil, concentrated oil, centrifuged essential oil, and juice oils [[12].12]. Centrifuged oil is classiﬁedclassified based on the extraction technology used [[11].11]. Oil type A is obtained viavia centrifugationcentrifugation ofof thethe oil/juiceoil/juice emulsion produced by screwing thethe wholewhole fruit in a press that crushescrushes thethe fruit.fruit. The traditionaltraditional method to extract essential oils is by cold pressing the citrus peels. The oil is presen presentt in oil sacs or oil glands located at different depths in the peel and cuticles of the fruit. Peel and cuticle oils are removed mechanically

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by cold pressing. Since cold pressing yields a watery emulsion, this emulsion is then centrifuged to separate out the essential oil [13]. Oil type B is obtained by rasping the peel to release the oil [11]. Because of its high solubility and pronounced ﬂavor characteristics, distilled essential oil is preferred for the manufacture of beverages [14], both alcoholic and non-alcoholic, especially cola beverages. The production of essential oil on a commercial scale is carried out using traditional methods such as steam distillation [6]. This kind of distillation is performed in batches. High temperatures are used for distillation, and the energetic consumption makes this process expensive. Moreover, high temperatures could induce degradation during distillation. Lime oil distilled from crushed lime fruit is rich in terpene alcohols and low in aldehydes and bicyclic hydrocarbons compared to pressed lime oil. During the production of distilled lime oil, acid-catalyzed reactions of bicyclic hydrocarbons occur, increasing the content of terpene alcohols, especially α-terpineol. Terpene aldehydes are almost completely lost during distillation [9]. These conventional methods show some disadvantages related to high energy costs and long extraction times [6]. During distillation, the citrus peels are exposed to boiling water or steam, releasing their essential oils through evaporation. As steam and essential oil vapors are condensed, both are collected and separated in a vessel traditionally called the “Florentine ﬂask” [13]. Distillation from lemon peel provides an oil yield of 0.21% from the total peel weight compared to 0.05% when using cold pressing [1].

1.4. Innovative Technologies To reduce the use of extractive solvents and the required energy, novel techniques are now available, including the use of supercritical extraction steam explosion, accelerated extraction at a high temperature and pressure, micro-waves, and ultrasound-assisted extraction [15]. The “Green Chemistry” concept is encouraging the development of these new en- vironmentally friendly techniques [15]. Although the above technologies have several advantages, they also require more sophisticated levels of technology that entail higher investment costs. These costs may not be easily attainable at industrial levels for average small- and medium-sized companies that extract essential oils in underdeveloped or moderately developed countries. Hence, the industry requires the development of alternative technologies with reduced energy consumption based, e.g., on the basic principle of steam entrainment distillation, which can be incorporated quickly and easily in industrial processes either through in- corporation into or the adaptation of existing technologies. This methodology has impli- cations for business success since the possible replacement of existing technology entails a sometimes-radical paradigm shift involving large investments of time and money. In this sense, continuous steam entrainment distillation could be a technological option for traditional steam entrainment processes.

1.5. Continuous Distillation by Steam The fractionation of essential oils rich in functional groups is a challenge with great prospects for the food and pharmaceutical industries because of the diverse range of medicinal properties belonging to these oils. To this end, the Centro de Investigación y Asistencia en Tecnología y Diseño del Estado de Jalisco developed a patented continuous steam distillation process that can be used to obtain essential oil in a more economical form than the traditional process of batch distillation [16] (Figure2). Continuous distillation is an economic process that presents opportunities and chal- lenges for the fractionation of essential oils. Research on continuous distillation was previously focused on the steam distillation of essential oils from a Mexican lime lab prototype at the level of 3 L [17] and recently from the rectiﬁcation of a tequila pilot facility at the level of 60 L [18]. Molecules 2021, 26, 4172 4 of 18

The continuous steam distillation process has many advantages and beneﬁts over the batch distillation process. For example, vapor consumption in continuous steam distillation saves up to 50% energy over batch distillation, and a potential fractionation of essential oil can be obtained. The residual juice can then be used to formulate a hydrating drink. These advantages result in a better oil quality because, in traditional distillation, the juice’s Molecules 2021, 26, x long contact times at high temperatures and acidic pH levels cause the degradation4 of of 19

oxygenated compounds [14].

FigureFigure 2. 2.Multifunctional Multifunctional distillation distillation prototype. prototype.

1.6. ContinuousContinuous Distillation distillation Process is an of economic Essential Oilsprocess that presents opportunities and chal- lengesThe for operational the fractionation procedure of essential for the continuous oils. distillation equipment is as follows: The juice flowResearch is fed on (F), continuous passing successively distillation throughwas previously each of thefocused five stages,on the thussteam obtaining distillation a residueof essential flow (W).oils from In each a Mexican of the stages, lime alab steam prototype stream at (S) the is fedlevel through of 3 L a[17] diffuser, and recently which risesfrom in the the rectification form of bubbles, of a tequila in which pilot the facility distilled at essential the level oil of emulsified60 L [18]. in the lemon juice is diffused.The continuous Then, the steam obtained distillation steam (V) process is condensed has many in advantages each of the and condensers benefits ofover each the stage,batch therebydistillation obtaining process. the Fo distillater example, stream vapor (D), consumption where the distilledin continuous oil is separatedsteam distilla- via decantationtion saves up for to subsequent 50% energy physicochemical over batch distillation, and chromatographic and a potential analysis. fractionation of essential oilThe can liquid be obtained. to be distilled The residual is fed (F) juice in the can first then stage be used (Figure to 3formulate) and travels a hydrating through eachdrink. Molecules 2021, 26, x ofThese the five advantages stages of result the equipment in a better oil until quality obtaining because, the in waste traditional stream distillation, (W). In each the5 of juice’s of the 19 steps,long bothcontact heat times exchangers at high carrytemperatures out evaporation and acidic of the pH volatiles, levels cause which the are degradation recovered in of eachoxygenated of the five compounds condensers [14]. to obtain the distillate (D).

1.6. Continuous Distillation Process of Essential Oils The operational procedure for the continuous distillation equipment is as follows: The juice flow is fed (F), passing successively through each of the five stages, thus obtaining a residue flow (W). In each of the stages, a steam stream (S) is fed through a diffuser, which rises in the form of bubbles, in which the distilled essential oil emulsified in the lemon juice is diffused. Then, the obtained steam (V) is condensed in each of the condensers of each stage, thereby obtaining the distillate stream (D), where the distilled oil is separated via decantation for subsequent physicochemical and chromatographic analysis. The liquid to be distilled is fed (F) in the first stage (Figure 3) and travels through each of the five stages of the equipment until obtaining the waste stream (W). In each of the steps, both heat exchangers carry out evaporation of the volatiles, which are recovered in each of the five condensers to obtain the distillate (D). This equipment has a heat recovery system that allows the recovery of the heat of the waste (W) from preheating the feed (F), which achieves additional energy savings when recovering the sensible heat. In each of the condensers, a distilled fraction (D) is obtained, which is then characterized. According to the desired profile or composition, this fraction (D) can be mixed [18]. Figure 3. Schematic ﬁgure of distillation. Figure 3.The Schematic aim of thisfigure work of distillation. was to validate a continuous steam distillation process (in terms of recuperation and energy efficiency) and to characterize the volatile fractions of essential 2. PersianResults lime and oilDiscussion both physically and chemically. 2.1. Content of Essential Oil in Citrus Juice The fruit of Persian citrus (Citrus latifolia Tanaka) contained 52% juice and 0.984 ± 0.287% v/v essential oil content. The essential oil recovered in the batch process was 94.7% ± 2.34, whereas that recovered through continuous distillation was 96.14% ± 2.32.

2.2. Efficiency of the Process and Steam Consumption in Continuous Distillation The continuous distillation process presented high oil recovery efficiency (up to 90%) and lower steam consumption (Figure 4) compared to traditional batch process distillation since steam consumption ranged from 32 to 60% for different steam levels (D/F).

100 90 80 70 60 50 40 30 20 10 0 % Energetic Comsume vs traditional % Energetic 0.2 0.4 0.6 Distillated vs Feed (D/F)

Figure 4. Percentage of steam consumption in continuous vs. traditional processes.

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This equipment has a heat recovery system that allows the recovery of the heat of the waste (W) from preheating the feed (F), which achieves additional energy savings when recovering the sensible heat. In each of the condensers, a distilled fraction (D) is obtained, which is then characterized. According to the desired proﬁle or composition, this fraction (D) can be mixed [18]. The aim of this work was to validate a continuous steam distillation process (in terms

of recuperation and energy efﬁciency) and to characterize the volatile fractions of essential PersianFigure 3. lime Schematic oil both figure physically of distillation. and chemically.

2.2. Results Results and and Discussion Discussion 2.1.2.1. Content Content of of Essential Essential Oil Oil in in Citrus Citrus Juice Juice TheThe fruit fruit of of Persian Persian citrus citrus (Citrus (Citrus latifolia latifoliaTanaka) Tanaka) contained contained 52% juice 52% and juice 0.984 and± 0.287%0.984 ± v/v0.287%essential v/v essential oil content. oil Thecontent. essential The essential oil recovered oil recovered in the batch in the process batch wasprocess 94.7% was± 94.7%2.34, whereas± 2.34, whereas that recovered that recovered through through continuous continuous distillation distillation was 96.14% was± 96.14%2.32. ± 2.32. 2.2. Efﬁciency of the Process and Steam Consumption in Continuous Distillation 2.2. Efficiency of the Process and Steam Consumption in Continuous Distillation The continuous distillation process presented high oil recovery efﬁciency (up to 90%) The continuous distillation process presented high oil recovery efficiency (up to 90%) and lower steam consumption (Figure4) compared to traditional batch process distillation and lower steam consumption (Figure 4) compared to traditional batch process distillation since steam consumption ranged from 32 to 60% for different steam levels (D/F). since steam consumption ranged from 32 to 60% for different steam levels (D/F).

100 90 80 70 60 50 40 30 20 10 0 % Energetic Comsume vs traditional % Energetic 0.2 0.4 0.6 Distillated vs Feed (D/F)

Figure 4. Percentage of steam consumption in continuous vs. traditional processes. Figure 4. Percentage of steam consumption in continuous vs. traditional processes.

2.3.Physicochemical Analysis The variance analysis showed only the fraction number to have a signiﬁcant effect on the refractive index, density, and optical rotation, whereas the distillate vs. feed (D/F) showed no signiﬁcant effect (Table1).

Table 1. Effect of the D/F and the fraction number in the refractive index, density, and optical rotation.

Density Optical Rotation Refractive Index Main (g/mL) (◦) Effects p-Value p-Value p-Value D/F 0.0003 0.4452 0.1354 Fraction number 0.0000 0.0004 0.0185 Molecules 2021, 26, 4172 6 of 18

The density and refractive index proﬁles showed an increase, whereas the optical Molecules 2021, 26, x rotation proﬁle showed a decrease, in the fractions distilled at the three7 dosage of 19 levels of

steam (Figure5).

Figure 5. Physical properties: (a) refractive index, (b) density, and (c) optical rotation. The results Figure 5. Physical properties: (a) refractive index, (b) density, and (c) optical rotation. The results ± are the averageare of the three average replicates of three ± standard replicates deviation.standard Different deviation. letters Different over a column letters indicate over a column indicate statistical differencesstatistical (Tukey, differences p < 0.05) (Tukey, p < 0.05).

With 95% confidence, no statistically significant differences were found in the physicochemical parameters (refractive index, optical rotation, and density) between fractions 1, 2, 3, 4, and 5 and the oil obtained by batch distillation. Compared to the reference, the

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The increase in the refractive index density (from 1.4693 to 1.47015) of the distilled fractions (from 0.8525 to 0.8585 g/mL) indicated a change in the composition of those fractions. Whereas F1 contained lighter compounds (terpenes), F5 contained heavier compounds (oxygenated). On the other hand, the observed decrease in optical rotation was linked to a reduction in the content of d-limonene, resulting in average values of +53.25◦ up to +46.2◦ for a D/F of 0.2 (Table2).

Table 2. Reference values for the physicochemical analysis.

Continuous Process Physicochemical Analysis Batch Process Fraction 3, D/F = 0.2 Refractive index at 20 ◦C 1.469 ± 0.0001 1.469 ± 0.00056 Optical rotation (◦) 48.875 ± 6.0 45.65 ± 1.6263 Density (g/mL) 0.857 ± 0.0021 0.860 ± 0.00777

2.3.1. Effect of the Fraction Number on the Density and Refractive Index of Essential Oil We observed significant differences (p < 0.05) in the refractive index and density of essential oil depending on the fraction number. Fractions were grouped by refractive index into four groups. The first group corresponded to Fraction 1, the second group included Fractions 2 and 3, and the third group included Fractions 3 and 4. Finally, fraction four included Fractions 4 and 5. The refractive index of the reference reported in the literature is 1.4751–1.4743 [19]. These values are higher than those obtained by the fractions (Figure5a), which may be because the residence time we explored was four hours, whereas the traditional process uses eight hours of contact time between the juice and the steam under acidic conditions, thereby impacting differences in the composition. For density (Figure5b), there were two homogeneous groups. The first (group a) included Fraction 1 and the second (group b) included Fractions 2, 3, 4, and 5. The density of the reference reported in the literature [19] is 0.8579–0.8556 g/mL whose range of values was close to that of Fraction 3 (Table2). For optical rotation, there were two homogeneous groups (Figure5c. The first (group a) included Fractions 1, 2, and 3, and the second (group b) included Fractions 2, 3, 4, and 5. The optical rotation of the reference reported in the literature (Table2) is +50.52 ◦–+46.84◦, whose range of values was close to that of Fraction 3. Usually, commercially available distilled essential oil presents similar physical–chemical properties (in terms of density and optical rotation) to those of F3 (Table2). With 95% confidence, no statistically significant differences were found in the physicochemical parameters (refractive index, optical rotation, and density) between fractions 1, 2, 3, 4, and 5 and the oil obtained by batch distillation. Compared to the reference, the only parameter that showed significant differences was the refractive index (Table2). According to Dugo et al. [11], the refractive index of Persian lime essential oil obtained by cold pressing was 1.4818, whereas the optical rotation was 47.3 degrees. The Persian lime essential oil obtained by hydro-distillation had similar values for the refractive index, the optical rotation, and the density than cold pressing or the fractions obtained by continuous steam distillation [14,19]. These results suggest that thermal treatment does not change the physicochemical properties of essential oils but possibly the composition. However, it is necessary to conduct further studies varying the residence time (4 h) of oil inside the continuous steam distillatory system over its physicochemical characteristics.

2.3.2. Effect of D/F on Density, Optical Rotation, and Density The refractive index and the optical rotation presented statistically significant differences with 95% confidence because of the D/F effect. For density and optical rotation, no statistically significant differences were found because of the D/F relationship. For the refractive index (Figure6), we observed two homogeneous groups: the first homoge- Molecules 2021, 26, x 8 of 19

only parameter that showed significant differences was the refractive index (Table 2). Ac- cording to Dugo et al. [11], the refractive index of Persian lime essential oil obtained by cold pressing was 1.4818, whereas the optical rotation was 47.3 degrees. The Persian lime essential oil obtained by hydro-distillation had similar values for the refractive index, the optical rotation, and the density than cold pressing or the fractions obtained by continuous steam distillation [14,19]. These results suggest that thermal treatment does not change the physicochemical properties of essential oils but possibly the composition. However, it is necessary to conduct further studies varying the residence time (4 h) of oil inside the continuous steam distillatory system over its physicochemical characteristics.

2.3.2. Effect of D/F on Density, Optical Rotation, and Density The refractive index and the optical rotation presented statistically significant differ- Molecules 2021, 26, 4172 8 of 18 ences with 95% confidence because of the D/F effect. For density and optical rotation, no statistically significant differences were found because of the D/F relationship. For the refractive index (Figure 6), we observed two homogeneous groups: the first homogeneous groupneous (group group a) included (group a) D/F included ratios D/Fof 0.2 ratios and 0.6, of 0.2 and and the 0.6, second and the group second (group group b) (group in- b) cludedincluded a D/F ratio a D/F of 0.4. ratio of 0.4.

FigureFigure 6. Refractive 6. Refractive Index vs. Index D/F. vs.The D/F.results The are results the average are the of averagethree replicates of three ± replicatesstandard devia-± standard tion. Differentdeviation. letters Different over lettersa column over indicate a column statistical indicate differences statistical differences(Tukey, p < (Tukey,0.05). p < 0.05).

2.4. Gas2.4. Chromatography Gas Chromatography for Key for Compounds Key Compounds AnalysisAnalysis by gas by chromatography gas chromatography permitted permitted to identify to identify an average an average of 47 compounds, of 47 compounds, eithereither in the in batch-distilled the batch-distilled essential essential oil or oilin orthe in fractions the fractions obtained obtained by continuous by continuous steam steam distillation.distillation. Usually, Usually, the number the number of compounds of compounds that can that be can identified be identified in citrus in citrus essential essential oil variesoil varies among among 30 compounds 30 compounds in citrus in citrus essential essential oil obtained oil obtained by hydro-distillation by hydro-distillation [13]. [13]. SimasSimas et al. et[20] al. reported [20] reported a total a totalof 38 ofcompou 38 compoundsnds in citrus in citrus essential essential oil also oil obtained also obtained by by hydro-distillation. hydro-distillation. On the other hand, Dugo et al. [11] reported 66 compounds in industrial cold-pressed On the other hand, Dugo et al. [11] reported 66 compounds in industrial cold-pressed essential oil of Persian lime. The higher number of compounds identified could be related essential oil of Persian lime. The higher number of compounds identified could be related to thermal treatment. Moreover, these authors obtained a higher number of compounds to thermal treatment. Moreover, these authors obtained a higher number of compounds in fractionated Persian lime essential oil. However, the fractions were obtained by glass in fractionated Persian lime essential oil. However, the fractions were obtained by glass column chromatography [11]. Although more compounds can be obtained by this method, column chromatography[11]. Although more compounds can be obtained by this method, it is more time-consuming compared to continuous steam distillation. it is more time-consuming compared to continuous steam distillation. No matter the method used to obtain essential oils from citrus species, the most No matter the method used to obtain essential oils from citrus species, the most abun- abundant compound is d-limonene. Other abundant compounds are β-pinene, geranial, dant compound is d-limonene. Other abundant compounds are β-pinene, geranial, neral, neral, p-cymene, and α-terpineol, which contribute significantly to the final composition of p-cymene, and α-terpineol, which contribute significantly to the final composition of es- essential oil [9,13,20]. Taking advantage of the abundance of these compounds, they were sentialused oil [9,13,20]. to analyze Taking the effect advantage of continuous of the steamabundance distillation of these on compounds, the composition they of were essential used oilto analyze fractions. the effect of continuous steam distillation on the composition of essential oil fractions,The D/F experimental ratio and fraction number had a very significant effect on the chemical profiles of the distilled fractions with 95% confidence, as shown in Table3 for d-limonene, p-cymene, α-terpineol, β-pinene, neral, and geranial. For γ-terpinene, only the factor of the fraction number was significant at 95%, whereas the effect of the D/F relationship was not significant. Consequently, the chemical composition was significantly different in the distilled fractions (Table3).

Table 3. Effect of the D/F ratio on the contents of d-limonene, γ-terpinene, p-cymene, α-terpineol, and β-pinene in the distilled fractions.

Main d-Limonene γ-Terpinene p-Cymene α-Terpineol β-Pinene Neral Geranial Effects p-Value p-Value p-Value p-Value p-Value p-Value p-Value D/F 0.0000 0.7087 0.0000 0.0000 0.0000 0.0000 0.0000 Fraction number 0.0005 0.0001 0.0027 0.0000 0.0000 0.0000 0.0000 Molecules 2021, 26, 4172 9 of 18

2.4.1. Effect of the Fraction Number on Relative Abundance The d-limonene, citral (neral + geranial), and β-pinene showed a decrease in composition (% of area) depending on the number fraction (Figures 7, 10, 12 and 13). Conversely, the p-cymene and α-terpineol concentrations showed an increase as the distillation time progressed (Figures 9 and 11). D-limonene is the most abundant and volatile component in lime essential oil. There- fore, d-limonene was the most abundant component in the vapor phases of all fractions obtained and was present in the greatest abundance during the first phase. The composition of d-limonene tended to reduce to approximately the fraction number, from an average content of 59.28% in the first fraction to an average content of 53.12% in the fifth fraction (Figure7). In this way, Fractions 1 and 2 featured higher d-limonene content than that obtained by the batch process, whose content was similar to that of Fraction 3. Conversely, Fractions 4 and 5 presented lower d-limonene content, with average values of 54.41% and 53.12%, respectively, in percentage of area. In the multiple range test, three homogeneous Molecules 2021, 26, x groups were obtained: a (containing F1), b (containing F2 and F3), and c (containing F3,10 of F4, 19

and F5).

a b bc cd d

FigureFigure 7. 7.Abundance Abundance proﬁle profile vs. vs. number number fraction fraction for for d-limonene. d-limonene. The The results results are are the the average average of threeof three replicates ± standard deviation. Different letters over a column indicate statistical differences replicates ± standard deviation. Different letters over a column indicate statistical differences (Tukey, (Tukey, p < 0.05). p < 0.05).

For γ-terpinene, the second most abundant terpene, no signiﬁcant differences were found in its abundance because of the effect of the fraction number. This is an interesting result given that γ-terpinene is chemically transformed to p-cymene during steam distillation and that p-cymene increasesa itsa abundancea as a functiona of thea fraction number. γ-terpinene may not have decreased in concentration because of its γ-terpinene range. On the one hand, γ-terpinene concentrations were much higher than those of p-cymene. However, this abundance was mainly attributable to the fact that γ-terpinene was produced during distillation from d-limonene, which is a d-limonene isomer and thus the most abundant component observed in the oil (Figure8). α-terpineol, one of the representative alcohols in essential oil, showed an increase in abundance relative to the fraction number (Figure9). This occurred because, as reported in the literature [14], α-terpineol is obtained through the hydration reactions of terpenes. During the production of distilled lime oil, acid-catalyzed reactions of bicyclic hydrocarbons occur, thus increasing terpene alcohols, especially α-terpineol.

Figure 8. Abundance profile vs. number fraction for γ-terpinene. The results are the average of three replicates ± standard deviation. Different letters over a column indicate statistical differences (Tukey, p < 0.05).

α-terpineol, one of the representative alcohols in essential oil, showed an increase in abundance relative to the fraction number (Figure 9). This occurred because, as reported in the literature [14], α-terpineol is obtained through the hydration reactions of terpenes. During the production of distilled lime oil, acid-catalyzed reactions of bicyclic hydrocarbons occur, thus increasing terpene alcohols, especially α-terpineol.

Molecules 2021, 26, x 10 of 19

a b bc cd d

Molecules 2021, 26, 4172 Figure 7. Abundance profile vs. number fraction for d-limonene. The results are the average10 of of 18 three replicates ± standard deviation. Different letters over a column indicate statistical differences (Tukey, p < 0.05).

a a a a a

FigureFigure 8. 8.Abundance Abundance proﬁle profile vs. vs. number number fraction fraction for forγ-terpinene. γ-terpinene. The The results results are are the the average average of threeof three replicates ± standard deviation. Different letters over a column indicate statistical differences Molecules 2021, 26, x replicates ± standard deviation. Different letters over a column indicate statistical differences (Tukey,11 of 19 (Tukey, p < 0.05). p < 0.05). α-terpineol, one of the representative alcohols in essential oil, showed an increase in abundance relative to the fractionα number (Figure 9). This occurred because, as reported in the literature [14], α-terpineol is-Terpineol obtained through the hydration reactions of terpenes. During10.0 the production of distilled lime oil, acid-catalyzed reactions of bicyclic hydrocarbons occur, thus increasing terpene alcohols, especiallycd α-terpineol. d 8.0 c b 6.0 D/F=0.6 a

% Area 4.0 D/F=0.4

2.0 D/F=0.2

0.0 F1 F2 F3 F4 F5 Fraction Number

FigureFigure 9. 9.Abundance Abundance proﬁle profile vs. vs. number number fraction fraction for forα -terpineol.α-terpineol. The The results results are are the the average average of of three replicatesthree replicates± standard ± standard deviation. deviation. Different Different letters overletters a columnover a column indicate indicate statistical statistical differences differences (Tukey, p(Tukey,< 0.05). p < 0.05).

TheTheβ β-pinene-pinene of of the the group group of terpenesof terpenes showed show theed same the behaviorsame behavior as d-limonene, as d-limonene, which decreasedwhich decreased in abundance in abundance with respect with respect to the fractionto the fraction number number (Figure (Figure 10). This 10). decrease This de- increaseβ-pinene in β-pinene was likely was due likely to thedue transformation to the transformation of bicyclic of bicyclic monoterpenes monoterpenes to monocyclic to mon- monoterpenesocyclic monoterpenes during heating. during heating. p-Cymene, which indicates the degree of transformation or degradation of the essential oil product due to contact with steam under acidic conditions at high temperatures, showed an increase relative to the fraction numberβ-Pinene (Figure 11) because of the dehydrogenation of cyclohexene rings of d-limonene or cyclohexadiene from γ-terpinene [9,14]. 8.0 a 7.0 6.0 5.0 b 4.0 D/F=0.6

% Area c 3.0 d D/F=0.4 d 2.0 D/F=0.2 1.0 0.0 F1 F2 F3 F4 F5 Fraction Number

Figure 10. Abundance profile vs. number fraction for β-pinene. The results are the average of three replicates ± standard deviation. Different letters over a column indicate statistical differences (Tukey, p < 0.05).

p-Cymene, which indicates the degree of transformation or degradation of the essential oil product due to contact with steam under acidic conditions at high temperatures, showed an increase relative to the fraction number (Figure 11) because of the dehydrogenation of cyclohexene rings of d-limonene or cyclohexadiene from γ-terpinene [9,14].

Molecules 2021, 26, x 11 of 19

α-Terpineol 10.0 cd d

8.0 c b 6.0 D/F=0.6 a

% Area 4.0 D/F=0.4

2.0 D/F=0.2

0.0 F1 F2 F3 F4 F5 Fraction Number

Figure 9. Abundance profile vs. number fraction for α-terpineol. The results are the average of three replicates ± standard deviation. Different letters over a column indicate statistical differences (Tukey, p < 0.05).

The β-pinene of the group of terpenes showed the same behavior as d-limonene, Molecules 2021, 26, 4172 which decreased in abundance with respect to the fraction number (Figure 10). This11 of de- 18 crease in β-pinene was likely due to the transformation of bicyclic monoterpenes to monocyclic monoterpenes during heating.

β-Pinene 8.0 a 7.0 6.0 5.0 b 4.0 D/F=0.6

% Area c 3.0 d D/F=0.4 d 2.0 D/F=0.2 1.0 0.0 Molecules 2021, 26, x 12 of 19 F1 F2 F3 F4 F5 Fraction Number

p-Cymene is produced via the degradation of γ-terpinene and indicates the oxidation FigureFigure 10. 10.Abundance Abundance proﬁle profile vs. vs. number number fraction fraction for forβ β-pinene.-pinene. The The results results are are the the average average of of three three replicatesand degradation ± standard processes deviation. of Different the essential letters oilover [1 4].a column Therefore, indicate in thisstatistical study, differences the p-cymene replicates ± standard deviation. Different letters over a column indicate statistical differences (Tukey, (Tukey,concentration p < 0.05). increased as the dosage of steam increased under acidic pH conditions. For pthe< 0.05). first fractions, p-cymene presented lower contents in Fractions 2, 3, 4, and 5. p-Cymene, which indicates the degree of transformation or degradation of the essential oil product due to contact with steam under acidic conditions at high temperatures, showed an increase relative to thep-Cymene fraction number (Figure 11) because of the dehydrogenation of cyclohexene rings of d-limonene or cyclohexadiene from γ-terpinene [9,14]. 4.5 4.0 3.5 3.0 c 2.5 b bc D/F=0.6 2.0 ab

% Area D/F=0.4 1.5 a 1.0 D/F=0.2 0.5 0.0 F1 F2 F3 F4 F5 Fraction Number

FigureFigure 11. 11.Abundance Abundance proﬁle profile vs. vs. number number fraction fraction for for p-cymene. p-cymene. The The results results are are the the average average of of three three replicates ± standard deviation. Different letters over a column indicate statistical differences replicates ± standard deviation. Different letters over a column indicate statistical differences (Tukey, (Tukey, p < 0.05). p < 0.05).

p-CymeneNeral and isgeranial, produced in viaaddition the degradation to showing of significantγ-terpinene differences and indicates between the oxidationthe differ- andent degradationfractions, decreased processes in of abundance the essential relative oil [14 to]. Therefore,the fraction in number. this study, The the first p-cymene fraction concentrationshowed the highest increased content, as the which dosage was of steam greate increasedr than that under from acidic batch pH distillation, conditions. whose For thecontent first fractions, was similar p-cymene to that of presented Fraction lower3. Neral contents and geranial in Fractions decreased 2, 3, because 4, and 5. the terpene aldehydesNeral and were geranial, almost incompletely addition to lost showing during significant distillation differences [9]. between the different fractions,The decreasedcitral content in abundance (neral + geranial), relative to which the fraction is shown number. in Table The 4, first agreed fraction with showed that re- theported highest in the content, literature which by was other greater authors than [8,16]. that fromThe citral batch content distillation, in the whose first fraction content wasfrom similarcontinuous to that distillation of Fraction (Figures 3. Neral 12 and and geranial 13) was decreased around 0.95% because of the the area, terpene and aldehydes that in the werefifth almostfraction completely was around lost 0.30% during [8]. distillation It was previously [9]. reported that the citral content in the Persian lime Citrus latifolia ranges from 0.5% to 2.2%. The value obtained through microwave-assisted hydrodistillation was 0.66% area, distributed among the isomers neral (Z-citral), with 0.29% area, and geranial (E-citral), with 0.38% area [16]. In this way, Frac- tions 1 and 2 presented a greater quantity of citral than that obtained by microwave-assisted hydro-distillation, whereas Fraction 3 presented a similar quantity of citral to that in the essential oil of Persian lime obtained by batch distillation. No statistically significant differences were found between the experimental runs carried out via batch distillation. (Table 4).

Table 4. Content of citral isomers (neral and geranial) in Persian lime.

Microwave-Assisted This Study’s This Study’s Batch Process [8] Hydrodistillation Compound Continuous Processes Batch Process (% Area) [6] (% Area) (% Area) (% Area) Neral 0.3 0.554 ± 0.240 Neral+ geranial: 0.29 Geranial 0.6 0.438 ± 0.611 0.5 to 2.2 0.38

Molecules 2021, 26, 4172 12 of 18

The citral content (neral + geranial), which is shown in Table4, agreed with that reported in the literature by other authors [8,16]. The citral content in the first fraction from continuous distillation (Figures 12 and 13) was around 0.95% of the area, and that in the fifth fraction was around 0.30% [8]. It was previously reported that the citral content in the Persian lime Citrus latifolia ranges from 0.5% to 2.2%. The value obtained through microwave-assisted hydrodistillation was 0.66% area, distributed among the isomers neral (Z-citral), with 0.29% area, and geranial (E-citral), with 0.38% area [16]. In this way, Fractions 1 and 2 presented a greater quantity of citral than that obtained by microwave- assisted hydro-distillation, whereas Fraction 3 presented a similar quantity of citral to that in the essential oil of Persian lime obtained by batch distillation. No statistically significant differences were found between the experimental runs carried out via batch distillation. (Table4).

Table 4. Content of citral isomers (neral and geranial) in Persian lime.

This Study’s This Study’s Microwave-Assisted Batch Process [8] Compound Continuous Processes Batch Process Hydrodistillation [6] (% Area) (% Area) (% Area) (% Area) Neral 0.3 0.554 ± 0.240 0.29 Molecules 2021, 26, x Neral+ geranial: 13 of 19 Geranial 0.6 0.438 ± 0.6110.5 to 2.2 0.38

Figure 12. Abundance profile vs. number fraction for neral. The results are the average of three Figure 12. Abundance proﬁle vs. number fraction for neral. The results are the average of three replicates ± standard deviation. Different letters over a column indicate statistical differences replicates ± standard deviation. Different letters over a column indicate statistical differences (Tukey, (Tukey, p < 0.05). p < 0.05).

The fractions presented signiﬁcant differences in the composition of minority compounds able to affect the aroma of essential oil fractions. Further studies are required at other residence times to evaluate the effects of these compounds on composition.

2.4.2. Effect of D/F on the Relative Abundance of Compounds The D/F had an effect on d-limonene, with statistically significant differences observed at 95% confidence in the abundance of the seven monitored compounds because of the effect of the D/F ratio. D-limonene, γ-terpinene, α-terpineol, neral, and geranial were presented as two homogeneous groups. The first homogeneous group (group a) included a D/F ratio of

Figure 13. Abundance profile vs. number fraction for geranial. The results are the average of three replicates ± standard deviation. Different letters over a column indicate statistical differences (Tukey, p < 0.05).

The fractions presented significant differences in the composition of minority compounds able to affect the aroma of essential oil fractions. Further studies are required at other residence times to evaluate the effects of these compounds on composition.

2.4.2. Effect of D/F on the Relative Abundance of Compounds The D/F had an effect on d-limonene, with statistically significant differences observed at 95% confidence in the abundance of the seven monitored compounds because of the effect of the D/F ratio. D-limonene, γ-terpinene, α-terpineol, neral, and geranial were presented as two homogeneous groups. The first homogeneous group (group a) included a D/F ratio of 0.2,

Molecules 2021, 26, x 13 of 19

Molecules 2021, 26, 4172 13 of 18

0.2,Figure and 12. the Abundance second homogeneousprofile vs. number group fraction (group for ne b)ral. included The results D/F are ratiosthe average of 0.4 of andthree 0.6 replicates ± standard deviation. Different letters over a column indicate statistical differences (Figure 14). (Tukey, p < 0.05).

Figure 13. Abundance profile vs. number fraction for geranial. The results are the average of three Figure 13. Abundance profile vs. number fraction for geranial. The results are the average of three replicates ± standard deviation. Different letters over a column indicate statistical differences replicates ± standard deviation. Different letters over a column indicate statistical differences (Tukey, (Tukey, p < 0.05). p < 0.05). The fractions presented significant differences in the composition of minority com- On the other hand, the γ-terpinene was presented as two homogeneous groups (Figurepounds 14 able). The to affect first group the aroma (group of a)essential included oil D/F fractions. ratios Further of 0.2 and studies 0.6, and are therequired second at groupother (groupresidence b) includedtimes to evaluate a D/F ratio the ofeffe 0.4.cts of these compounds on composition. Based on the homogeneous groups obtained in the statistical analysis, the highest relative2.4.2. Effect abundances of D/F on (% the area) Relative of neral Abundance and geranial, of Compounds d-limonene, β-pinene, and α-terpineol were reachedThe D/F with had the an D/Feffect 0.4 on and d-limonene, 0.6 relationships, with statistically with no differences significant observed differences between ob- theseserved two at conditions.95% confidence Therefore, in the the abundance process condition of the seven that obtainedmonitored the compounds greatest amount because of citralof the with effect a reductionof the D/F in ratio. steam consumption was a D/F of 0.4. WithD-limonene, 95% confidence, γ-terpinene, no statisticallyα-terpineol, significantneral, and geranial differences were were presented found as in two the F1ho- fractionsmogeneous with groups. respect The to the first experimental homogeneous runs group carried (group out in a) batch included distillation a D/F (Tableratio of5). 0.2,

Table 5. Composition of essential oil of Persian lemon by different processes.

This Study’s Continuous This Study’s Microwave-Assisted Processes Fraction 1 Batch Process [8] Compound Batch Process Hydrodistillation [6] D/F = 0.4 (% Area) (% Area) (% Area) (% Area) d-limonene 60.09 ± 1.21 55.26 ± 1.75 59.016 61.93 γ-terpinene 14.87 ± 0.58 15.14 ± 0.38 16.089 16.93 α-terpineol 2.76 ± 0.60 2.78 ± 034 2.145 0.39 β-pinene 6.57 ± 0.55 8.17 ± 0.58 6.035 11.35 p-cymene 0.64 ± 0.26 0.64 ± 0.07 1.507 0.21 neral 0.34 ± 0.007 0.55 ± 0.24 0.091 0.29 geranial 0.68 ± 0.17 0.44 ± 0.61 0.121 0.38

Since some fractions presented concentrations close to those of distilled oil obtained via batch processes, this process was explored under a residence time of four hours, leading to a reduction greater than 50% compared to traditional processes. Furthermore, the juice efﬂuent can be used for the preparation of a rehydrating drink. However, other process conditions must be explored to study the effects of residence time on the composition and Molecules 2021, 26, 4172 14 of 18

oil recovery efﬁciency. The results suggest that a longer process time may be desirable, as the liquid and vapor contact time increased under acidic conditions and high temperatures. Molecules 2021, 26, x 15 of 19 This high-temperature, acidic environment favored the transformation of compounds, thereby impacting the ﬁnal composition of the distilled oil similar to traditional processes.

FigureFigure 14.14.( a(a)) d-limonene, d-limonene, ( b()bα) -terpineol,α-terpineol, (c )(cγ) -terpinene,γ-terpinene, (d )(d neral,) neral, (e) ( geranial,e) geranial, (f) β(f-pinene,) β-pinene, (g) p-cymene.(g) p-cymene. The The results results are theare averagethe average of three of three replicates replicates± standard ± standard deviation. deviation. Lowercase Lowercase letters letters indicate indicate homogenous homogenous groups. groups.

With 95% confidence, no statistically significant differences were found in the F1 fractions with respect to the experimental runs carried out in batch distillation (Table 5).

Molecules 2021, 26, 4172 15 of 18

In the ﬁrst stage, it is possible to recover the essential oil contained in the juice. Although it is possible to obtain fractions with different compositions to the studied compounds, more studies are needed to obtain well-differentiated terpene, sesquiterpene, or oxygenated fractions. Greater separation could be achieved by using packed columns in the experimental distillation process. This phenomenon will be explored in future works.

3. Methodology 3.1. Process 3.1.1. Raw Material The raw material used for the experimental tests was Persian lime (Citrus latifolia Tanaka) from the region of San Martin Hidalgo, Jalisco. Fresh, green-looking Persian lime juice was used in the initial stages of yellowing and was obtained by pressing the whole fruit using a screw press. We ﬁrst ensured that the fruit was well developed, green in color, and had no signs of rot. Each lime was manually washed with water to eliminate any foreign agents. The lime was then processed in a helical press in the pilot plant of CIATEJ. This press has a conical shape and features perforated walls where the juice comes out. The fruit was crushed between these walls to break the cells containing the essential oil, forming a juice–oil emulsion.

3.1.2. Batch Distillation by Steam The juice–oil emulsion was then transported to the batch distillation equipment described above, where steam was injected to increase the temperature, thus evaporating the water and essential oil. The vapors resulting from the distillation passed through a coil condenser where they changed into a liquid form. This liquid formed by the water and essential oil was deposited into Florentine vessels, where the essential oil was separated by decantation from the water in which it was carried. The recovered essential oil was packaged in amber glass jars, where a small amount of sodium sulfate was added to eliminate any trace of moisture. The amber glass jars containing the essential oil were then stored in refrigeration as a ﬁnished product for further analysis.

3.1.3. Continuous Distillation by Steam Continuous distillation was carried out using the distillation equipment previously described. Tests were performed in triplicate for each of the experimental stages under the same conditions, which are described later in the experimental design.

3.1.4. Experimental Design for Continuous Distillation The experimental design was unifactorial 31, where the factor was the ratio of distilled steam flow to feed flow (D/F), with the following levels: 0.2, 0.4, and 0.6. Response Variables The response variables evaluated were as follows: 1. The oil recovery efficiency (%); 2. The percentage of steam consumption compared to the traditional process (%). The steam consumption was determined using a steam flux Vortex Sensor Prowirl 200 (Endress and Hauser) to quantify the amount of steam consumed during distillation (Magnetic Flow Transmitter Meter, Endress and Hauser).

3.2. Physical-Chemical Analysis of the Distilled Essential Oil Once the distillation tests were carried out, both oil fractions obtained from the continuous tests were subjected to the following physicochemical analyses: the determination of the relative density, the determination of the refractive index, and the determination of the rotary power (optical rotation). We obtained these values to determine the oil’s characteristics and compare those characteristics with those of the oil obtained via the traditional method of steam entrainment. Molecules 2021, 26, 4172 16 of 18

3.2.1. Determination of Relative Density The relative density was determined based on the French Standard NF T 75-111 in accordance with the international standard [21,22], as published by the International Organization for Standardization. For this analysis, a DMA 35N digital densimeter was used under the conditions speciﬁed in the standard.

3.2.2. Determination of the Refractive Index The refractive index was determined based on the French Standard NF T 75-112 in accordance with the international standard [20], as published by the International Organization for Standardization. A Leica Mark II Plus Abbe Refractometer was used for this analysis.

3.2.3. Determination of the Rotatory Power To determine the rotatory power, a Poli Science Model SR6 polarimeter was used. This analysis was performed based on the French Standard NF V 75-113 in accordance with the international standard [21], as published by the International Organization for Standardization.

3.2.4. Determination of the Content of Essential Oil in Lime Juice As in previous analyses, the content of volatile oils in the juice was determined for the raw material according to French Standard NF V 03-409 in technical agreement with the international standard ISO/DIS 6571.

3.2.5. Determination of the Essential Oil Components by Gas Chromatography The volatile composition of the essential oil or fractions was determined according to the norm AFNOR [21]. Oils in the distilled fractions were analyzed by gas chromatography (GC) using an HP 6890 gas chromatograph equipped with a flame ionization detector (FID). Separation was performed on an HP-1 methyl siloxane capillary column (50 m × 0.2 mm i.d. × 0.33 µm film thickness) with helium as the carrier gas (0.8 mL/min). The oven temperature was programmed initially at 75 ◦C and was then increased at a rate of 2.5 ◦C/min to 240 ◦C. A volume of 0.1 µL was injected at 1:150 split ratio. The injection port and detector temperatures were 260 and 280 ◦C, respectively. A mixture of air (400 mL/min), hydrogen (40 mL/min), and nitrogen as an auxiliary gas (10 mL/min) was also fed to the detector. The compounds of interest were identified by the retention time from the injection of pure standards (SIGMA-ALDRICH, purity ≥ 95%) under the same conditions as the sample. The concentration of the compounds was expressed in the % area. Each fraction was analyzed in duplicate.

4. Conclusions The process of continuous steam distillation to obtain essential oil from Persian lime juice presented the highest oil recovery efﬁciencies (up to 90%) alongside lower steam consumption compared to traditional batch process distillation (up to 50%). More studies are being done to evaluate the application of continuous steam distillation to other essential oils, such as the one from citrus peels. The chemical composition of the oil fractions obtained via continuous distillation showed signiﬁcant differences for each major component based on the effect of the D/F ratio and the number of the fraction. The chemical compounds that presented the greatest variation in concentration throughout the various fractions were as follows: d-limonene, p-cymene, β-pinene, α-terpineol, and citral (neral + geranial).

Author Contributions: Conceptualization, J.D.P.-d.l.R., M.E.-E. and G.G.-M.; formal analysis, J.D.P.- d.l.R., A.C.-M., G.G.-M. and E.A.-G.; investigation, J.D.P.-d.l.R., M.D.M.-A. and J.R.G.-H.; methodology, J.D.P.-d.l.R. and M.E.-E.; project administration, J.D.P.-d.l.R.; resources, J.D.P.-d.l.R.; supervision, J.D.P.-d.l.R., M.E.-E. and E.A.-G.; validation, M.D.M.-A. and J.R.G.-H.; visualization, M.D.M.-A. and J.R.G.-H.; writing—original draft, J.D.P.-d.l.R., M.E.-E. and A.C.-M.; writing—review and editing, Molecules 2021, 26, 4172 17 of 18

J.D.P.-d.l.R., M.E.-E. and G.G.-M. All authors have read and agreed to the published version of the manuscript. Funding: This research was funded by Fondo Mixto CONACYT and the Veracruz Government— Ignacio de la Llave, grant number 37480; by CONACYT Sectorial Ciencia Básica SEP-CONACYT No. A1-S-34273 and by COECYTJAL, grant number 9156-2020. Institutional Review Board Statement: Not applicable. Informed Consent Statement: Not applicable. Data Availability Statement: The data presented in this study are available on request from the corresponding author. Acknowledgments: Thanks to CIATEJ for the facilities and support during the development of the present work. Thanks to Abiel Alba, Ernesto Rodríguez; and Ignacio Orozco for their technical support in the pilot plant. Conﬂicts of Interest: The authors declare no conﬂict of interest. Sample Availability: Samples of the compounds are not available from the authors.

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