Long-Chain Alcohol-Modified Micellar Systems and Their Application

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Thermodynamics, Transport, and Fluid Mechanics

Long-chain alcohol-modified micellar systems and their application in a continuous extraction process Oliver Fellechner, Sebastian Rotzolk, and Irina Smirnova Ind. Eng. Chem. Res., Just Accepted Manuscript • DOI: 10.1021/acs.iecr.8b04617 • Publication Date (Web): 09 Jan 2019 Downloaded from http://pubs.acs.org on January 10, 2019

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Industrial & Engineering Chemistry Research

Long-chain alcohol-modified micellar systems and their application in a continuous extraction process Oliver Fellechner*, Sebastian Rotzolk, and Irina Smirnova. Hamburg University of Technology, Institute of Thermal Separation Processes, Eißendorfer Straße 38, D-21075 Hamburg, Germany

Abstract The integration of an in situ extraction into biocatalytic processes is often limited by the toxicity of organic solvents. Therefore, it is desirable to use water-based extraction systems, for example aqueous micellar two-phase systems. They can be used, for instance, for extraction of valuable products from microalgae cultures. Recently, the nonionic surfactant ROKAnol NL5 was identified as a suitable surfactant for this purpose, since it forms an upper micellar phase, enabling an easy separation of whole-cell biocatalysts. However, its application at temperatures below 45 °C is limited by unstable phase boundaries, whereas the maximal temperature to ensure the vitality of the most microalgae cultures is around 40 °C. To overcome this problem, addition of long-chain alcohols to the surfactant-water mixture during extraction is suggested in this work. Using 1-hexanol a continuous extraction process with the model solute trans-cinnamic acid at 40 °C in a stirred column could be realized. The results of a new suggested system

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Industrial & Engineering Chemistry Research 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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water/ROKAnol NL5/1-hexanol at 40 °C (extraction yield 𝑌𝑐𝑜𝑛𝑡. = 97.67 ± 0.14%, enrichment factor 𝑙𝑜𝑔10 𝑇𝐶𝐴 = 2.42 ± 0.03, number of theoretical stages 𝑁𝑡ℎ𝑒𝑜 = 4.45 ± 0.16) are comparable to the system water/ROKAnol NL5 at 45 °C (𝑌𝑐𝑜𝑛𝑡. = 99.26 ± 0.24%, 𝑙𝑜𝑔10 𝑇𝐶𝐴 = 2.60 ± 0.10, 𝑁𝑡ℎ𝑒𝑜 = 5.88 ± 0.67), ensuring, however, no damage of microalgae.

Keywords: Nonionic surfactants; Long-chain additives; Continuous extraction; Cloud point extraction

1. Introduction Due to an easy and fast cultivation, green microalgae are more and more in the focus of research activities1. One major benefit of algae over land-biomass like straw and wood is the absent lignin in the molecular structure. However, not only the conversion of microalgae in the biorefinery or an energetic use, but also isolation of their high-value products is of interest. Fermentation products from microalgae cultures range from fatty acids, lipids, and pigments to the production of substances for the cosmetic as well as for the food industry and pharmaceutical applications2,3. Moreover, microalgae can be used in the production of biodiesel4–6. The common process for the purification of microalgae products, as described in the literature so far, can be divided into four steps: Cell disruption, dehydration, product isolation and product purification7. These process steps are cost-intensive; also a cultivation step of microalgae cells has to be performed at the beginning for every batch. Therefore, the integration of a first separation step into the reaction systems was applied for the in situ extraction of microalgae products. Initially, Kleinegris et al.8 mentioned dodecane as a


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biocompatible solvent. Nevertheless, the authors mentioned a phase toxicity and an emulsion formation of dodecane9. Alternatively, Glembin et al.10–12 introduced aqueous micellar two-phase systems for the in situ extraction of hydrophobic fermentation products from microalgae cultures. Aqueous micellar two-phase systems are based on aqueous solutions with nonionic surfactants. After exceeding the critical micelle concentration (cmc), dynamic aggregates, so-called micelles, are formed. The hydrophobic tails are in the micellar core, surrounded by the hydrophilic heads, which interact with the aqueous bulk phase. Based on this effect, hydrophobic molecules (solutes), like fermentation products of microalgae cultures, can be concentrated in micelles. Aqueous micellar two-phase systems show a lower critical solution temperature: upon increasing the temperature, two liquid phases are formed. This temperature is called cloud point temperature (CPT). The system consists of an aqueous-rich (surfactant-lean) and a surfactant-rich phase. Due to an accumulation of surfactant in the surfactant-rich phase, also the hydrophobic solutes are accumulated in this phase13. The cloud point extraction finds various applications in literature. Examples are the extraction of wastewater compounds, extraction of dyes and compounds from microbial fermentation processes13–17. Trakultamupatam et al.13 mentioned the cloud point extraction for the wastewater treatment. Thereby, the nonionic surfactant Triton X-114 from Dow Chemical Inc. was used to extract phenol and toluene as aromatic tracer substances, to show the feasibility of the cloud point extraction in a rotating disc contactor. The results show that 87.5% of toluene was extracted in a continuous process. According to their publication, the limitation of the separation process and its scale-up is the entrainment of the surfactant-rich phase13.


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An application for biological systems is given by Dhamole et al.18, who worked on the in situ extraction of butanol from fermentative production. The problem they stated in fermentative production of butanol is the toxicity of the product on bacteria. Therefore, they identified Pluronic L62 as a biocompatible nonionic surfactant. The partition coefficient of butanol was for this system between three and four. The butanol production was increased by 225% by this mean18. Our group worked in recent years on the in situ extraction of hydrophobic products from microalgae broths. First, Glembin et al.10 showed the in situ extraction of hydrophobic compounds from microalgae products with the nonionic surfactants Triton X-114, Tergitol TMN 6 and Tergitol 15-S-7.

The

used

microalgae

cultures

were

Chlamydomonas reinhardtii,

Chlorella vulgaris, and Acutodesmus obliquus. They showed that the microalgae cells were accumulated in the aqueous phase, whereby fatty acids could be enriched in the micellar phase. Moreover, Acutodesmus obliquus showed a good biocompatibility with the nonionic surfactants. Based on these results, Racheva et al.19 demonstrated a continuous in situ extraction of microalgae compounds from Acutodesmus obliquus with the nonionic surfactant Triton X-114. Trans-cinnamic acid (CA) was used as a tracer substance to give quantitative statements for the extraction process. Thereby, a maximum yield of 79.1 ± 0.2% of trans-cinnamic acid was extracted. The partition coefficient was in this experiment log10PCA = 1.2 ± 0.2 at 40 °C. Nevertheless, one major drawback for the application with Triton X-114 is the fact, that the density of the micellar phase is higher as that of the aqueous phase, where the cells are present (ρsurfactant > ρwater). The sedimentation of the microalgae cells leads to a contamination of the extract with microalgae cells and an additional solid/liquid separation after the extraction process is necessary.


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With the introduction of ROKAnol NL5, which forms an upper micellar phase (ρsurfactant < ρwater), Racheva et al.20 avoided the sedimentation of microalgae cells into the micellar phase. Furthermore, ROKAnol NL5 is cosmetic-grade and the extracted microalgae compounds can be used as leave-in products for an application in the cosmetic industry. The partition coefficient of trans-cinnamic acid was determined to log10PCA = 1.2 ± 0.2 at 45 °C and showed an overall yield in a continuous process of nearly 100%. However, for many algae strains the temperatures higher than 40°C are hardly tolerable so that the extraction temperature should be decreased accordingly. Unfortunately the extraction system water/ROKAnol NL5 shows phase stability problems below 45 °C and thus can not be applied. At the same time it is known, that hydrophobic additives like alcohols, ethers and non-dissociated carboxylic acids influence the liquid-liquid equilibria. Thus, in this work we aim to evaluate different long chain alcohols as additives to water/ROKAnol NL5 systems in order to enable the extraction at 40°C. The ultimate goal thereby is to present a concept of an in situ extraction process for microalgae compounds, which allows a recycle stream of the microalgae culture to the fermenter and the use of the solvent as a leave-in product for further industrial applications. 2. Materials & Methods 2.1. Nonionic surfactant The nonionic surfactant ROKAnol NL5 from PCC Exol an ethoxylated alcohol (C9 – C11) is a clear liquid with an average molecular weight of 380 g mol-1 and has a hydrophilic-lipophilic balance of 11.6. According to the product data sheet, the cloud point of an aqueous solution is between 33 and 39 °C and the pure surfactant solution has a density of approximately 0.97 g ml -1 (25 °C). The critical micelle concentration of ROKAnol NL5 is, according to the manufacturer, 160 mg L-1. Moreover, the surfactant solution can contain low aliphatic alcohols.


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2.2. Additives The alcohols used in this work as additives are shown in Table 1. Table 1: List of used organic additives in this work with producer, purity, CAS-number, density and FEMA number. Food-grade or cosmetic-grade additives are marked with a (*). Data were taken from the safety data sheets provided by the producer. Name

Producer

Purity

CAS

Density [g cm-3] at 20 °C

FEMA number

1-hexanol

Merck KGaA

≥ 98

111-27-3

0.82

2567

trans-2-hexen-1-ol*

Alfa Aesar

≥ 96

928-95-0

0.843

2562

1-octanol

Alfa Aesar

≥ 99

111-87-5

0.835

2800

1-hexadecanol*

Alfa Aesar

≥ 98

36653-82-4 0.817

2554

2.3. Algae cultivation The microalgae culture Acutodesmus obliquus provided by SSC GmbH was cultivated in a 1 L Schott bottle at room temperature. The culture was gassed with an airflow of 1 L min-1 and additional 4-5 vol.-% of CO2. The gas flow ensured an axial dispersion and a magnetic stirrer approved the radial dispersion. A fertilizer (1 g L-1, Flory Basis Fertilizer 1, Euflor, Germany) and 1.6 g L-1 KNO3 (Carl Roth GmbH & Co. KG, Germany) were added to the cultivation broth. The pH was adjusted to pH 7 by the addition of 1 M sodium hydroxide (Th. Geyer GmbH & Co. KG, Germany) respectively 1 M HCl (Merck KGaA, Germany). The microalgae culture was illuminated with red LEDs, which were attached to the bottle. 2.4. Algae vitality The microalgae vitality was measured by pulse amplitude modulated (PAM) fluorometry. 15 mL of the cultivation media were added to a 100 mL beaker glass. The beaker glass was stored for 10


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min in darkness to ensure an ideal activity of the photoreceptors. To avoid a sedimentation of the cells, the microalgae broth was mixed from time to time. The relative photosynthetic activity RPA was calculated according to equation 1 and 2. 𝐹𝑉 𝐹𝑀

=

𝐹𝑀 − 𝐹0

(1)

𝐹𝑀 𝐹 ( 𝑉)

𝑅𝑃𝐴 =

Where

𝐹𝑀 𝑖 𝐹𝑉 ( ) 𝐹𝑀 𝑟𝑒𝑓

𝐹𝑉 𝐹𝑀

(2)

is the maximum quantum yield and can be calculated with the maximum 𝐹𝑀 and

minimum 𝐹0 fluorescence yield for a reference state ref and a sample i. 2.5. Heat resistance test The influence of the temperature on the microalgae vitality was tested in a range of 40 to 45 °C in steps of 1 °C. 15 mL samples were tempered in a water bath (‘DL 20 KP’, LAUDA DR. R. Wobser GmbH & Co. KG, Germany) at the corresponding temperature and the relative photosynthetic activity was measured every 60 min. The experiment was performed for 240 min. Each experiment was performed fourfold. 2.6. Toxicity test The effect of ROKAnol NL5 on the microalgae vitality was measured analog and in the same time interval as described for the heat resistance test. A ROKAnol NL5 concentration in a range of 0.44 to 2.62 g L-1 was added to the microalgae broth. The samples were stored at room temperature. 2.7. Measurement of the cloud point temperature The CPT was measured by the “cloud point method”, described by Racheva et al.20. Therefore, samples with an overall mass of 50 g were prepared with wROKAnol NL5 = 5 wt%, an additive molality


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up to 2.0 moladditive kg-1 and the rest was added as deionized water. The samples were solvent homogenized and tempered in a water bath (‘DL 20 KP’, LAUDA DR. R. Wobser GmbH & Co. KG, Germany) at 25 °C. The temperature was increased with a heating rate of 1 °C min-1 up to 45 °C. The CPT was determined visually. The temperature was measured indirect with a thermometer in a reference sample to avoid contamination of the samples. Every CPT was measured in triplicate. 2.8. Determination of partition coefficients Trans-cinnamic acid (CA, Sigma Aldrich, purity 99%) was chosen as a model substance for all extraction experiments. CA has a molecular weight of 148.16 g mol-1 and due to its hydrophobic character as well as low water solubility (0.23 ± 0.01 g L-1 at 25 °C), it is well known as a tracer substance for the cloud point extraction19,20. The partition coefficients were determined for samples with an overall mass of 14 g (5 wt.% ROKAnol NL5, wCA = 0.2 gCA kg-1 and changing the concentration of additives). As solvent additives, the alcohols 1-hexanol, tans-2-hexen-1-ol, 1-octanol, and 1-hexadecanol were used. The samples were mixed for 30 min in an overhead shaker and settled at 40 °C overnight. The phases were separated as described before. The logarithmic partition coefficient 𝑙𝑜𝑔10 Pαβ was calculated CA by the ratio between mass fraction of trans-cinnamic acid in micellar α and aqueous phase β in equation 3. 𝛼𝛽

𝑙𝑜𝑔10 𝑃𝐶𝐴 = 𝑙𝑜𝑔10

𝛼 𝑤𝐶𝐴

(3)

𝛽

𝑤𝐶𝐴

2.9. Continuous countercurrent extraction Continuous extraction experiments were performed in a stirred extraction column, built by Normag Labor- und Prozesstechnik GmbH. The column had an inner diameter of 30 mm and an active height of 1000 mm, divided into 32 stirring cells. The column was tempered 0.5 °C above


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the desired temperature of the experiment to balance the heat loss. The aqueous phase was introduced with a gear pump ISMATEC ISM901 (IDEX Health & Science GmbH, Germany). The solvent was a ROKAnol NL5 solution, containing a mass fraction of ROKAnol NL5 according to the LLE of each system at the corresponding temperature. The surfactant solution was pumped with a peristaltic pump ISMATEC MCP (IDEX Health & Science GmbH, Germany). All continuous experiments were performed in a countercurrent mode. Due to the densities of the phases, the surfactant solution was introduced at the bottom, whereas the heavy feed solution was pumped in at the head of the column. Samples of the raffinate and the extract were collected at the bottom and head of the column. The continuous experiments were evaluated by the mass fraction of ROKAnol NL5 in the 𝑅 raffinate 𝑤𝑅𝑂𝐾𝐴𝑛𝑜𝑙 𝑁𝐿5 , the enrichment factor 𝑇𝐶𝐴 (equation 4) and the extraction yield of CA 𝑌𝑐𝑜𝑛𝑡.

(equation 5). To measure the ROKAnol NL5 concentration in the raffinate, samples were partly evaporated in an oven (T 5060 E, Heraeus, Germany). 𝑇𝐶𝐴 =

𝐸 𝑤𝐶𝐴

(4)

𝑅 𝑤𝐶𝐴

𝑌𝑐𝑜𝑛𝑡. =

𝐸 𝑚̇𝐶𝐴 𝐹 𝑚̇𝐶𝐴

∙ 100% =

𝐹 𝑅 𝑚̇𝐶𝐴 −𝑚̇𝐶𝐴 𝐹 𝑚̇𝐶𝐴

∙ 100%

(5)

𝐸 𝑅 𝐸 𝑅 With 𝑤𝐶𝐴 and 𝑤𝐶𝐴 as the weight fractions of CA in the extract E and raffinate R and 𝑚̇𝐶𝐴 , 𝑚̇𝐶𝐴 𝐹 and 𝑚̇𝐶𝐴 are the corresponding CA mass flows of extract, raffinate and feed F.

The number of theoretical stages 𝑁𝑡ℎ𝑒𝑜 were calculated according to Sattler21 (equation 6).

𝑁𝑡ℎ𝑒𝑜 =

𝐹

𝑆

𝑤𝐶𝐴

𝑤𝐶𝐴

𝑤 𝑤𝐶𝐴 𝑙𝑛[(𝜀−1) ∙ ( 𝐶𝐴 𝑅 + 𝑅 ) +1]

𝑙𝑛(𝜀)

−1

(6)

With 𝜀 as extraction factor (equation 7); 𝛼𝛽

𝜀 = 𝑃𝐶𝐴 ∙

𝑚̇𝐸

(7)

𝑚̇𝑅


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𝜈 describes the feed-to-solvent ratio (equation 8). 𝜈=

𝑚̇𝐹

(8)

𝑚̇𝑆

2.10.

Analytics

High-performance liquid chromatography Trans-cinnamic acid was analyzed with an Agilent 1200 series reversed phase high-performance liquid chromatography (HPLC, Agilent Technology Inc., USA, degasser, quaternary pump, tempered autosampler, column thermostat, diode array detector DAD, refractive index detector RID). The column Eclipse XDB-C18, 5 µm, 4.6x150 mm with an additional Zorbax RX-C8 precolumn was used. CA was detected at 275 nm for high concentration (wCA > 2·10-2 wt%) and for lower concentration at 310 nm. ROKAnol NL5 was detected with the RID. As eluent, a mixture of 90 vol.-% acetonitrile and 10 vol.-% filtered and demineralized water was used. Density measurements Samples were prepared according to partition coefficient experiments and the phases were separated as described before. The densities were measured in an oscillating u-tube (DMA 4500 M, Anton Paar GmbH, Austria). The densities were measured according to the temperature of each experiment. Each density was determined in triplicate.

3. Results and Discussion In this work, the influence of long-chain alcohols on the extraction performance and cloud point temperature on the mixture ROKAnol NL5-water is studied with the purpose to design an efficient extraction process at temperatures below 40°C. The alcohols were chosen as additives also since


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they represent cosmetic-grade additives to guarantee an overall cosmetic-grade extraction process for relevant applications. 3.1. Extraction of microalgae compounds The microalgae culture Acutodesmus obliquus provided by SSC GmbH was used for the in situ extraction of microalgae products. First, a continuous extraction experiment was performed at 45°C (𝑚̇𝐹 = 6 mL min-1, 𝑚̇ 𝑆 = 2 mL min-1, stirrer speed 12 rpm, wROKAnol NL5= 16.80 wt%). At these conditions, the microalgae culture showed no vitality after the extraction process. Moreover, it was visually observed that ROKAnol NL5 extracted green pigments from the cells. To clarify this fact, the relative photosynthetic activity (RPA) at different temperatures and contact times was measured (Figure 1). Confirming with the results from the extraction experiment, the RPA decreased continuously over time. However, at temperatures lower than 40°C, no RPA loss is detected.

Figure 1: Relative photosynthetic activity (RPA) of Acutodesmus obliquus at different temperatures and contact times. The relative photosynthetic activity RPA was measured every


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hour. The RPA is given in a grey scale, whereby white symbolizes a RPA of around one. The maximum error in the RPA is ± 0.05. At the same time it was confirmed, that the surfactant itself had no significant negative influence on RPA as shown in Figure 2. Within the first two hours, the RPA decreases with increasing ROKAnol NL5 concentration and afterward a fluctuation of the RPA around a stable value is observed (in the tested time interval). However, the minimal RPA measured was 0.91, which was considered as satisfactory for extraction application, especially concerning the standard deviation of these measurements series (± 0.07).

Figure 2: Relative photosynthetic activity of Acutodesmus obliquus as a function of the ROKAnol NL5 concentration (in wt%) and time. The experiments were performed for 0.00, 0.04, 0.09, 0.17 and 0.26 wt% ROKAnol NL5 and were measured every hour at 25 °C. The relative photosynthetic activity is given in a grey scale. White is a RPA around one. The maximum error in RPA is ± 0.07 for this experimental series.


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Thus, it was suggested that the extraction process with ROKAnol NL5 can be realized at temperatures ≤ 40 °C. However, a decrease of the process temperature not only has an influence on microalgae vitality, but also influences the phase equilibria. By decreasing the process temperature from 45 °C to 40 °C, the ROKAnol NL5 concentration in the micellar phase decreases from wROKAnol NL5 = 16.75 wt% (own work, measured by HPLC) to wROKAnol NL5 = 11.90 wt% (Racheva et al.20, phase diagram). Moreover, no stable phase boundary during experiments in the continuous extraction column could be observed at 40°C. Comparable effects were described in the literature for the nonionic surfactant Tergitol 15-S-7 (ρTergitol 15-S-7 < ρWater)22. Thus, the simple decrease of the temperature does not solve the given extraction problem. Therefore, additives were tested to tune the liquid-liquid equilibrium and density differences of water/ROKAnol NL5 and to enable the extraction at these conditions. 3.2. Influence of organic hydrophobic additives on the extraction system In previous works it was shown, that the addition of low molecular weight compounds allows to adjust the cloud point extraction system by changing its physico-chemical properties. For instance, due to the addition of D-glucose (ρD-glucose > ρWater), the density difference in a system with an upper micellar phase was increased, due to unequal partitioning of D-glucose between the two 𝐴𝑃 𝑀𝑃 phases (𝑐𝐷−𝑔𝑙𝑢𝑐𝑜𝑠𝑒 > 𝑐𝐷−𝑔𝑙𝑢𝑐𝑜𝑠𝑒 )22. This led to a successful implementation of a system

water/Tergitol 15-S-7/D-glucose in a stirred extraction column22. However, the addition of sugars to microalgae cultures could lead to a contamination by bacteria or other microorganisms in an open fermentation process. Moreover, the overall mass of D-glucose required for an industrial process would be immense. To minimize the effect of an additive on the fermentation process, additives, which accumulate in the micellar phase, should be evaluated. In literature, organic compounds (e.g. aliphatic


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alcohols, hydrocarbons, acids, ester, ketones, and ethers) were examined with regards to their effect on the CPT as well as on shape and size of micelles23–26. However, information about the influence of additives on phase equilibria and especially on partition coefficients of solutes as well as on phase densities required for the extraction process are rarely available27. In this work, four alcohols (tans-2-hexen-1-ol, 1-hexanol, 1-octanol and 1-hexadecanol) were systematically examined as additives for micellar systems. Generally, alcohols with a C-chain up to C3 increase the CPT due to their high solubility in water (the longer the C-chain is, the higher should be its influence on the CPT) 23,24. Among available alcohols, trans-2-hexen-1-ol (FEMA number 2562) and 1-hexadecanol (FEMA number 2554) were chosen as food-grade and/or cosmetic-grade additives to maintain a cosmetic-grade process as established with ROKAnol NL520. In a first step, the influence of all alcohols on the process-relevant properties (CPTs, density differences, surfactant concentration) were compared. Finally, continuous experiments were performed with 1-hexanol as an additive. 3.3. Comparison of cloud point temperatures, density differences and ROKAnol NL5 concentration in micellar phase To compare the influence of different alcohols, samples with a surfactant concentration of wROKAnol NL5 = 5 wt% were prepared with addition of alcohols at different concentrations. The results are shown in Figure 3. Both alcohols with a C-chain of C6 show a similar influence on the CPT, whereas 1-octanol has, in accordance with the literature, a larger influence24. The effect of 1-hexadecanol is lower than 1-octanol and more comparable with 1-hexanol. Also, Díaz-Fernández et al.23 showed that there was no increase in the influence between heptan-1-ol and decan-1-ol. Obviously, the influence on the CPT is not linear within a homologues series of additives as thus is not a simple effect of solubility. Further it was observed that samples containing


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more than w1-octanol = 0.13 wt% tend to form an emulsion of the aqueous and micellar phase. The same problem occurred with 1-hexanol and trans-2-hexen-1-ol over walcohol = 0.29 wt% and for 1-hexadecanol at w1-hexadecanol = 0.16 wt%. Consequently, it was not possible to proceed further experiments above these concentrations.

Figure

3:

Cloud

point

temperatures

of

ROKAnol

NL5-water-alcohol

solutions

(wROKAnol NL5 = 5 wt% in all cases). In the most systems with alcohols, a stable phase boundary was formed at 40 °C and thus the phases could be separated and analyzed (Figure 4). With increasing concentration of all additives, the concentration of ROKAnol NL5 in the micellar phase increases. This trend is reciprocal to the trend of the CPT and can be explained by the shift of the LLE and a resulting higher equilibrium concentration of ROKAnol NL5 in the micellar phase.


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Figure 4: Concentration (weight percents) of ROKAnol NL5 in the micellar phase (total wROKAnol NL5 = 5 wt%) for the four alcohols trans-2-hexen-1-ol, 1-hexanol, 1-octanol and 1-hexadecanol. Data for a binary system ROKAnol NL5-water (no additive), taken from Racheva et al.20, is given for comparison. Further, the densities of both, the aqueous and the micellar phase, were measured at 40 °C. The results are shown in Figure 5. For all tested alcohols the density difference increases with an increasing additive concentration. This can be explained by two phenomena: first, the alcohols have a lower density than water (ρAlcohol < ρWater) and accumulate in the micellar phase (compare partition of alcohols with Mehling et al.27); secondly, the surfactant is concentrated in the micellar phase due to a decrease of the CPT and leads to an increase in the density difference (ρROKAnol NL5 < ρWater). By comparing the weight fractions of alcohol and surfactant in the system, it is apparent that the increase in the density difference is more influenced by the enrichment of ROKAnol NL5 in the micellar phase. The highest density difference reached is 3.15 ± 0.08 g L-1 (w1-hexanol = 0.30 wt%).


16

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Figure 5: Density differences of the aqueous and micellar phases of water-ROKAnol NL5 solutions upon addition of alcohols (trans-2-hexen-1-ol, 1-hexanol, 1-octanol and 1-hexadecanol). The initial concentration of ROKAnol NL5 was wROKAnol NL5 = 5 wt%. To highlight the benefit of organic long-chain additives for adjusting the phase equilibrium and physicochemical properties of micellar systems, the densities of the aqueous and micellar phase of a system water/ROKAnol NL5/1-hexanol are shown in Table 2. Table 2: Densities of aqueous (AP) and micellar phases (MP) at different 1-hexanol concentrations. The concentration of 1-hexanol refers to the initial monophasic solution, containing water, ROKAnol NL5, and 1-hexanol. w1-Hexanol [wt%]

ρAP [g L-1]

ρMP [g L-1]

0.00

992.68 ± 0.36

991.90 ± 0.02

0.12

992.65 ± 0.01

990.53 ± 0.04

0.18

992.67 ± 0.01

990.36 ± 0.01

0.21

992.60 ± 0.01

990.18 ± 0.00

0.24

992.65 ± 0.01

989.82 ± 0.01


17


0.30

992.60 ± 0.07

Page 18 of 34

989.45 ± 0.08

Table 2 shows clearly that the additives influence the density of the micellar phase, but not of the aqueous one. This is in agreement with the enrichment of long-chain alcohols in the micellar phase as discussed before. Influence of long-chain alcohols on the partition coefficients of trans-cinnamic acid (CA) For a further examination of the influence of long-chain additives on micellar systems, the partition coefficient of CA was measured at different alcohol concentrations (Table 3). With increasing additive concentration, the partition coefficient increases as well. At constant additive concentration the partition coefficient increases with increasing length of additives’ carbon chain (however, this effect is smaller). The highest partition coefficient of CA in the tested systems

was

achieved

at

the

highest

additive

concentration

(w1-hexanol = 0.29 wt%,

𝛼𝛽

𝑙𝑜𝑔10 𝑃𝐶𝐴 = 1.09 ± 0.00). Partitioning of non-charged solutes (as CA in its non-dissociated form) is dominated by hydrophobic interactions between the solute and the micelles hydrophobic core28–30. Due to the accumulation of the long chain alcohols in the micellar phase, which leads to the increase of its hydrophobicity, the partition coefficient of CA increases at higher alcohol concentration. Furthermore, the surfactant concentration in the micellar phase also increases upon alcohol addition, as shown before in Figure 4. This leads to a smaller volume of the micellar phase and thereby, to higher solute concentration. 𝛼𝛽

Table 3: Partition coefficients (𝑙𝑜𝑔10 𝑃𝐶𝐴 ) of trans-cinnamic acid in systems containing water/ROKAnol NL5/additive.

The

initial

ROKAnol NL5

concentration

was

wROKAnol NL5 = 5 wt%.


18

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𝛼𝛽

Additive

wAdditive [wt%]

Without additive

0

0.89 ± 0.01

0.09

0.95 ± 0.01

0.12

0.97 ± 0.01

0.15

1.03 ± 0.01

0.19

1.07 ± 0.01

0.24

1.05 ± 0.00

0.29

1.09 ± 0.00

0.13

0.94 ± 0.06

0.16

0.97 ± 0.01

0.20

0.98 ± 0.00

0.24

1.01 ± 0.01

0.29

1.02 ± 0.08

0.08

0.95 ± 0.01

0.12

0.98 ± 0.01

0.02

0.91 ± 0.01

0.04

0.95 ± 0.01

0.08

0.99 ± 0.01

0.13

1.00 ± 0.01

0.16

0.96 ± 0.01

1-hexanol

trans-2-hexen-1-ol

1-octanol

1-hexadecanol

Continuous

countercurrent

extraction

𝑙𝑜𝑔10 𝑃𝐶𝐴

with

an

extraction

system

water/ROKAnol NL5/1-hexanol As discussed above, without additives, a continuous extraction process with a system water/ROKAnol NL5 could not be realized at 40 °C, since the density difference between the aqueous and micellar phase was too low to give a stable phase boundary. The addition of long-


19


Page 20 of 34

chain additives improved the density difference, so it was assumed that it would enable the extraction. 1-hexanol was chosen as the most suitable additive. The extraction parameters were chosen according to the previously determined parameters (𝑚̇𝐹 = 6 mL min-1, 𝑚̇ 𝑆 = 2 mL min-1, stirrer speed 12 rpm) to allow a comparison between the process at 45 °C without additive and 40 °C with 1-hexanol (w1-hexanol = 1.01 wt% in micellar phase). The stirrer speed was however decreased to 10 rpm, since relatively small density differences (see Figure 5) and surface tensions (see e.g. Ritter et al.22) should lead to a sufficient residence time and droplet size even with a small energy input. The visually observed droplet sizes were in the same range in both experiments (12 and 10 rpm). The results of the extraction are shown in Figure 6. The concentration of CA in the raffinate is stable after 480 min, showing that the process reached a steady state. The concentration of ROKAnol NL5 in the raffinate are oscillating between wROKAnol NL5 = 0.2 and 0.3 what is comparable to results at 45 °C. Nevertheless, the CA concentration is still decreasing slowly after 600 min in extract, which can be explained by a time delay between samples from raffinate and extract. The extraction yield, enrichment factor and number of theoretical stages as determined in the process are shown in Table 4. With an active height of 1 m in the extraction column, the number of theoretical stages per meter is 4.45 at 40 °C and 5.88 at 45 °C.


20

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Figure 6: Concentration profiles of CA (in wt%) in raffinate and extract for continuous countercurrent extraction experiments at 40 °C (light grey) with 10 rpm and a 1-hexanol 𝑀𝑃 concentration in the initial micellar phase (solvent) of 𝑤1−ℎ𝑒𝑥𝑎𝑛𝑜𝑙 = 1.01 wt% and 45 °C (grey)

with 12 rpm without additive. Table 4: Process evaluation of the cloud point extraction of trans-cinnamic acid with ROKAnol NL5 (𝑌𝑐𝑜𝑛𝑡. : Extraction efficiency, 𝑇𝐶𝐴 : Enrichment factor, 𝑁𝑡ℎ𝑒𝑜 : Number of theoretical stages). Temperature [°C]

Stirrer [rpm]

40 45

speed

𝑌𝑐𝑜𝑛𝑡. [%]

𝑙𝑜𝑔10 𝑇𝐶𝐴 [-]

𝑁𝑡ℎ𝑒𝑜 [-]

10

97.67 ± 0.14

2.42 ± 0.03

4.45 ± 0.16

12

99.26 ± 0.24

2.60 ± 0.10

5.88 ± 0.67

Overall, satisfactory results were achieved for the extraction process at 40°C. It proves that the surfactant-water system can be successfully modified by using additives, such as long chain alcohols, so that extraction at low temperatures (T ≤ 40 °C) can be realized. This finding can help


21


Page 22 of 34

to further extend the applications of surfactant-based extraction processes for temperature-sensible systems. 4. Conclusion In this work, the application of long-chain alcohols as additives for a continuous surfactantbased extraction process was shown for the first time. The cosmetic-grade nonionic surfactant ROKAnol NL5 was used for this purpose. ROKAnol NL5 forms an upper micellar phase, which improves the separation of whole-cell biocatalysts like microalgae. The main drawback of ROKAnol NL5-water system, namely unstable phase boundary at low temperatures, could be overcome by addition of long-chain alcohols. With increasing alcohol concentration, the CPT decreases, whereas the concentration of ROKAnol NL5 in the micellar phase increases. Moreover, the density difference between the aqueous and micellar phase as well as the partition coefficient of CA increases, what is favorable for the extraction. Applying these finding, the system water/ROKAnol NL5/1-hexanol has been successfully processed in a continuous countercurrent extraction of microalgae cultures. In regard to the extraction efficiency, comparable results to the system without additives were achieved (𝑌𝑐𝑜𝑛𝑡. = 97.67 ± 0.14%, 𝑙𝑜𝑔10 𝑇𝐶𝐴 = 2.42 ± 0.03, 𝑁𝑡ℎ𝑒𝑜 = 4.45 ± 0.16), but algae damage could be avoided. The substitution of 1-hexanol with a cosmetic-grade alcohol like trans-2-hexen-1-ol could ensure an overall cosmetic-grade process to use the solvent water/ROKAnol NL5/alcohol as a leave-in product for the cosmetic industry. Corresponding Author *Tel.: +49 40 428784073. Fax: +49 40 428784072. E-mail: [email protected] Author Contributions


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The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. Funding Sources The authors appreciate the financial support of DFG (project SM 82/14-1). Notes ACKNOWLEDGMENT The authors appreciate the financial support of DFG (project SM 82/14-1). ABBREVIATIONS 𝑐𝑖𝑥 , concentration of component i in phase x; CA, trans-cinnamic acid; cmc, critical micelle concentration; CPT, cloud point temperature; E, extract; F, feed; 𝐹0 , minimum fluorescence 𝐹

yield; 𝐹𝑀 , maximum fluorescence yield; 𝐹 𝑉 , maximum quantum yield; FEMA, Flavor Extract 𝑀

𝑦

Manufactures Association; HPLC, high performance liquid chromatography; 𝑚̇𝑖 , mass flow of 𝑥𝑦

component i in stream y; 𝑁𝑡ℎ𝑒𝑜 , number of theoretical stages; 𝑃𝑖 , partition coefficient of component i between the phases x and y; R, raffinate; RPA: relative photosynthetic activity; S, solvent; 𝑇𝑖 , enrichment factor of component i; 𝑤𝑖𝑥 , weight fraction of component i in phase x; 𝑌𝑐𝑜𝑛𝑡. , extraction yield; ε, extraction factor; ν, feed-to-solvent ratio; ρ, density REFERENCES (1)

Barclay, W.; Apt, K.; Dong, X. D. Commercial Production of Microalgae via Fermentation. In Handbook of Microalgal Culture; Wiley-Blackwell, 2013; pp 134–145.

(2)

Bai, M.-D.; Cheng, C.-H.; Wan, H.-M.; Lin, Y.-H. Microalgal Pigments Potential as Byproducts in Lipid Production. J. Taiwan Inst. Chem. Eng. 2011, 42 (5), 783–786.


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(3)

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Falaise, C.; François, C.; Travers, M.-A.; Morga, B.; Haure, J.; Tremblay, R.; Turcotte, F.; Pasetto, P.; Gastineau, R.; Hardivillier, Y.; Leignel, V.; Mouget, J.-L. Antimicrobial Compounds from Eukaryotic Microalgae against Human Pathogens and Diseases in Aquaculture. Mar. Drugs 2016, 14 (9), 159.

(4)

Bellinger, E. G.; Sigee, D. C. Introduction to Freshwater Algae. In Freshwater Algae; Wiley-Blackwell, 2015; pp 1–42.

(5)

Menetrez, M. Y. An Overview of Algae Biofuel Production and Potential Environmental Impact. Environ. Sci. Technol. 2012, 46 (13), 7073–7085.

(6)

Chisti, Y. Biodiesel from Microalgae Beats Bioethanol. Trends Biotechnol. 2008, 26 (3), 126–131.

(7)

Grima, E. M.; Fernández, F. G. A.; Medina, A. R. Downstream Processing of Cell Mass and Products. In Handbook of Microalgal Culture; Wiley-Blackwell, 2013; pp 267–309.

(8)

Kleinegris, D. M. M.; Janssen, M.; Brandenburg, W. A.; Wijffels, R. H. Two-Phase Systems: Potential for in Situ Extraction of Microalgal Products. Biotechnol. Adv. 2011, 29 (5), 502–507.

(9)

Kleinegris, D. M. M.; Es, M. A. van; Janssen, M.; Brandenburg, W. A.; Wijffels, R. H. Phase Toxicity of Dodecane on the Microalga Dunaliella Salina. J. Appl. Phycol. 2011, 23 (6), 949–958.

(10) Ingram, T.; Storm, S.; Glembin, P.; Bendt, S.; Huber, D.; Mehling, T.; Smirnova, I. Aqueous Surfactant Two-Phase Systems for the Continuous Countercurrent Cloud Point Extraction. Chem. Ing. Tech. 2012, 84 (6), 840–848.


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Page 25 of 34 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(11) Glembin, P.; Racheva, R.; Kerner, M.; Smirnova, I. Micelle Mediated Extraction of Fatty Acids from Microalgae Cultures: Implementation for Outdoor Cultivation. Sep. Purif. Technol. 2014, 135, 127–134. (12) Glembin, P.; Kerner, M.; Smirnova, I. Cloud Point Extraction of Microalgae Cultures. Sep. Purif. Technol. 2013, 103, 21–27. (13) Trakultamupatam, P.; Scamehorn, J. F.; Osuwan, S. Scaling Up Cloud Point Extraction of Aromatic Contaminants from Wastewater in a Continuous Rotating Disk Contactor. I. Effect of Disk Rotation Speed and Wastewater to Surfactant Ratio. Sep. Sci. Technol. 2005, 39 (3), 479–499. (14) Appusamy, A.; John, I.; Ponnusamy, K.; Ramalingam, A. Removal of Crystal Violet Dye from Aqueous Solution Using Triton X-114 Surfactant via Cloud Point Extraction. Eng. Sci. Technol. Int. J. 2014, 17 (3), 137–144. (15) Appusamy, A.; Purushothaman, P.; Ponnusamy, K.; Ramalingam, A. Separation of Methylene Blue Dye from Aqueous Solution Using Triton X-114 Surfactant. J. Thermodyn. 2014, 2014, 1–16. (16) Purkait, M. K.; DasGupta, S.; De, S. Performance of TX-100 and TX-114 for the Separation of Chrysoidine Dye Using Cloud Point Extraction. J. Hazard. Mater. 2006, 137 (2), 827– 835. (17) Wang, Z.; Xu, J.-H.; Chen, D. Whole Cell Microbial Transformation in Cloud Point System. J. Ind. Microbiol. Biotechnol. 2008, 35 (7), 645–656. (18) Dhamole, P. B.; Wang, Z.; Liu, Y.; Wang, B.; Feng, H. Extractive Fermentation with NonIonic Surfactants to Enhance Butanol Production. Biomass Bioenergy 2012, 40, 112–119.


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(19) Racheva, R.; Tietgens, N.; Kerner, M.; Smirnova, I. In Situ Continuous Countercurrent Cloud Point Extraction of Microalgae Cultures. Sep. Purif. Technol. 2018, 190, 268–277. (20) Racheva, R.; Rahlf, A. F.; Wenzel, D.; Müller, C.; Kerner, M.; Luinstra, G. A.; Smirnova, I. Aqueous Food-Grade and Cosmetic-Grade Surfactant Systems for the Continuous Countercurrent Cloud Point Extraction. Sep. Purif. Technol. 2018, 202, 76–85. (21) Sattler, K. Thermische Trennverfahren: Grundlagen, Auslegung, Apparate; John Wiley & Sons, 2012. (22) Ritter, E.; Racheva, R.; Jakobtorweihen, S.; Smirnova, I. Influence of D-glucose as additive on thermodynamics and physical properties of aqueous surfactant two-phase systems for the continuous micellar extraction. Chem. Eng. Res. Des. 2017, 121, 149–162. (23) Díaz-Fernández, Y.; Rodríguez-Calvo, S.; Pérez-Gramatges, A. Influence of Organic Additives on the Cloud Point of PONPE-7.5. Phys Chem Chem Phys 2002, 4 (20), 5004– 5006. (24) Gu, T.; Galera-Gómez, P. A. The Effect of Different Alcohols and Other Polar Organic Additives on the Cloud Point of Triton X-100 in Water. Colloids Surf. Physicochem. Eng. Asp. 1999, 147 (3), 365–370. (25) Rocha, S. A. N.; Costa, C. R.; Celino, J. J.; Teixeira, L. S. G. Effect of Additives on the Cloud Point of the Octylphenol Ethoxylate (30EO) Nonionic Surfactant. J. Surfactants Deterg. 2013, 16 (3), 299–303. (26) Sharma, R.; Bahadur, P. Effect of Different Additives on the Cloud Point of a Polyethylene Oxide-Polypropylene Oxide-Polyethylene Oxide Block Copolymer in Aqueous Solution. J. Surfactants Deterg. 2002, 5 (3), 263–268.


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(27) Mehling, T.; Ingram, T.; Smirnova, I. Experimental Methods and Prediction with COSMORS to Determine Partition Coefficients in Complex Surfactant Systems. Langmuir 2012, 28 (1), 118–124. (28) Materna, K.; Szymanowski, J. Separation of Phenols from Aqueous Micellar Solutions by Cloud Point Extraction. J. Colloid Interface Sci. 2002, 255 (1), 195–201. (29) Sosa Ferrera, Z.; Padrón Sanz, C.; Mahugo Santana, C.; Santana Rodrı́guez, J. J. The Use of Micellar Systems in the Extraction and Pre-Concentration of Organic Pollutants in Environmental Samples. TrAC Trends Anal. Chem. 2004, 23 (7), 469–479. (30) Liu, C.-L.; Nikas, Y. J.; Blankschtein, D. Novel Bioseparations Using Two-Phase Aqueous Micellar Systems. Biotechnol. Bioeng. 1996, 52 (2), 185–192.


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For Table of Contents Only


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Figure 1: Relative photosynthetic activity (RPA) of Acutodesmus obliquus at different temperatures and contact times. The relative photosynthetic activity RPA was measured every hour. The RPA is given in a grey scale, whereby white color symbolizes a RPA of around one. The maximum error in the RPA is ± 0.05. 129x80mm (300 x 300 DPI)



Figure 2: Relative photosynthetic activty of Acutodesmus obliquus as a function of the ROKAnol NL5 concentration (in wt%) and time. The experiments were performed for 0.00, 0.04, 0.09, 0.17 and 0.26 wt% ROKAnol NL5 and were measured every hour at 25 °C. The Relative photosynthetic activity is given in a grey scale. White is a RPA around one. The maximum error in RPA is ± 0.07 for this experimental series. 129x80mm (300 x 300 DPI)


Page 30 of 34

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Figure 3: Cloud point temperatures of ROKAnol NL5-water-alcohol solutions (wROKAnol NL5 = 5 wt% in all cases). 149x80mm (300 x 300 DPI)



Figure 4: Concentration (weight percents) of ROKAnol NL5 in micellar phase (total (wROKAnol NL5 = 5 wt%) for the four alcohols trans2-hexen-1-ol, 1-hexanol, 1-octanol and 1-hexadecanol. Data for a binary system ROKAnol NL5-water (no additive) taken from Racheva et al.20 is given for comparison. 149x80mm (300 x 300 DPI)


Page 32 of 34

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Figure 5: Density differences of the aqueous and micellar phases of water-ROKAnol NL5 solutions upon addition of alcohols (trans-2-hexen-1-ol, 1-hexanol, 1-octanol and 1-hexadecanol). The initial concentration of ROKAnol NL5 was wROKAnol NL5 = 5 wt%. 149x80mm (300 x 300 DPI)



Figure 6: Concentration profiles of CA (in wt%) in raffinate and extract for continuous countercurrent extraction experiments at 40 °C (light grey) with 10 rpm and a 1-hexanol concentration in the initial micellar phase (solvent) of w1-hexanolMP = 1.01 wt% and 45 °C (grey) with 12 rpm without additive. 149x80mm (300 x 300 DPI)


Page 34 of 34

Long-Chain Alcohol-Modified Micellar Systems and Their Application

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