Abstract
British Phycological APPLIED PHYCOLOGY Society 2023, VOL. 4, NO. 1, 1–14 Understanding and using algae https://doi.org/10.1080/26388081.2022.2125829 Growth comparison of Arthrospira platensis in different vessels: standard cylinder vs. enhanced surface area at low light a b b a Jonathan L. Gal , Nolan R. Cole , Dennis L. Eggett and Steven M. Johnson a b Department of Microbiology and Molecular Biology, College of Life Sciences, Brigham Young University, Provo, UT, USA; Department of Statistics, College of Physical and Mathematical Sciences, Brigham Young University, Provo, UT, USA ABSTRACT ARTICLE HISTORY Received 14 March 2022 A laboratory scale prototype of a novel tube called AlgaTube™ (patent pending) was evaluated for Accepted 9 September 2022 growth of Arthrospira platensis (commonly referred to as Spirulina) in a controlled, 10-day batch trial at low light intensity. The two cultures (control and prototype) were grown in identical KEYWORDS conditions inside a closed incubator space at 32°C with a mean light intensity of 55 µmol AlgaTube™; Arthrospira; −2 −1 photons m s LED light, consisting mostly of red and blue wavelengths with some full spectrum biomass; cyanobacteria; −1 background radiation. OD was measured daily and converted to Concentration (C , g l ) using feed; fertilizer; nutrition; 590 d a standard curve. Four independent replicates of the trial were run. Six biomass growth metrics photobioreactor; wastewater; Spirulina were evaluated and compared. Across the board, all six growth metrics showed higher rates of biomass growth in the AlgaTube™. Maximum concentration was 34% higher (p = 0.02), cumulative production was 41% higher (p = 0.02), specific growth rate was 17% higher (p = 0.02), maximum specific growth rate was 23% higher (p = 0.02), mean daily production was 41% higher (p = 0.001), and maximum daily production was 38% higher (p = 0.19). We conclude that the novel shape of the AlgaTube™ prototype increased biomass growth rates. We believe that this study constitutes successful proof-of-concept for the AlgaTube™, but further studies are needed to optimize its performance. Introduction Industrial applications can be divided into three Photosynthetic microorganisms have been studied as steps: (1) growth and production, (2) harvest and pro- a feedstock for many commercial and industrial appli- cessing, and (3) marketing and delivery. The present cations, including: CO sequestration, biofuels, human study is focused exclusively on the production step, nutrition, fish and livestock feed, air purification, ferti- which is common to all the applications listed above. lizer, cosmetics, pharmaceutical manufacturing, waste- Thus, the present study (and the associated product) water remediation, and bioplastics (Stanley & Jones, could have a broad impact on all of the applications 1976; Benemann, 1979; Kumar, Dasgupta, Nayak, mentioned above. It is a platform technology. Lindblad, & Das, 2011; Priyadarshani & Biswajit, 2012; Production technology has historically been focused Rasala & Mayfield, 2015; Christaki, Karatzia, & Florou- on the open pond system, also known as the “raceway”, Paneri, 2010; Poonam & Sharma, 2017; Adeniyi, and some still use that technique (Cyanotech). More Azimov, & Burluka, 2018; Joshi, 2018; Liu, Pemberton, recently, closed production systems have been built Lewis, Scales, & Martin, 2020; Arora, Kumar, Bose, Li, & using clear, cylindrical piping (Phytobloom). These are Kulshrestha, 2021; Chong et al., 2021; Shahi et al., 2022). generally referred to as tubular photobioreactors The United States Department of Energy investigated (tPBRs). Closed systems are more costly to build, but photosynthetic microorganisms as a feedstock for bio- they offer advantages such as protection against con- fuels during the 1980s and 1990s (Sheehan, Dunahay, tamination, control of culture conditions, and increased Benemann, Roessler, & Weissman, 1998). Several com- production rates (Clippinger & Davis, 2019). panies are currently marketing such products [e.g., Different variables can impact the production rate of Cyanotech, TrueAlgae, and Algeternal] (Tacon & photobioreactors (PBRs). Temperature, light, pH, sali- Metian, 2008). During 2019, the US National nity, nutrients, and gas exchange can all be varied. Each Renewable Energy Laboratory published a review of variable can have a distinct impact on the culture growth the state-of-the-art for photosynthetic algae production rate (Hoseini, Almodares, Afsharzadeh, Shahriari, & (Clippinger & Davis, 2019). Montazeri, 2014; Kendirlioglu & Cetin, 2017; Olaizola & CONTACT Steven M. Johnson stevenj@byu.edu © 2023 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. 2 J. L. GAL ET AL. Duerr, 1990; Soni, Sudhakar, & Rana, 2017, 2019; tubes that are widely used as state-of-the-art in tubular Srinivasan & Illanjiam, 2021a, 2021b; Tayebati, Shariati, photobioreactors. The novel shaped culture vessel tested Soltani, & Tehrani, 2020; Uslu et al., 2009; Wang, Fu, & here was shown to significantly improve the growth Liu, 2007). rate. The increase was most likely due to changing the Cultures of photosynthetic algae and cyanobacteria shape of the sidewalls of the cylindrical culture vessel have been observed to experience growth inhibition by significantly reducing the self-shading effect by increas- the shadow effect, also referred to as the self-shading ing the surface area of the sidewalls of the culture vessel. effect (Frontasyeva et al., 2009; Olaizola & Duerr, 1990; This new configuration may help to improve the eco- Soni, Sudhakar, & Rana, 2019). This occurs when the nomics of any industrial process that utilizes photosyn- cell density is high enough that the cells shade each thetic microorganisms such as cyanobacteria and algae. other, thus blocking each other’s access to needed light. At particularly high concentrations, light penetra- Materials and methods tion is severely restricted. (González-Camejo et al., 2019; Shigesada & Okubo, 1981). Culture vessels Other studies have successfully shown that increasing The surface area of the sidewalls of the AlgaTube™ was the surface-area-to-volume ratio (SA/V) improves the the primary variable that we isolated and evaluated in growth rate by exposing greater numbers of cells to this experiment. Here, we use the term Photon Surface direct irradiance, thus reducing the shadow effect Area (PSA) to label this key variable and to differenti - (Converti, Lodi, Del Borghi, & Solisio, 2006; da Silva ate it from the cross-sectional surface area (CSSA), et al., 2016; Fremento et al., 2013). In those studies, SA/ which is a different measure altogether. In the present V was increased by reducing the diameter of cylindrical −1 study, PSA Vol is equivalent to the “SA/V” metric tubes, an effect that is inherent in the Euclidean geome- used by Converti, da Silva and other referenced try of standard cylindrical tubes. In the present study, authors. The control flask was a standard, cylinder- a different approach was taken to increasing the SA −1 −1 shaped vessel. Both vessels were custom made by 3- Vol ratio of culture vessels. The SA Vol was chan- D printing, using the same resin and the same manu- ged, not by reducing the diameter, but by using corru- facturing technology (DMS Somos® Watershed XC gated sidewalls in lieu of conventional sidewalls. 11,122 resin and Stereolithography (SLA)). The other The purpose of the present study was to evaluate dimensions of the two flasks were chosen such that the a prototypic tube with enhanced, corrugated surface remaining flasks were chosen such that the remaining area, the AlgaTube™, for its ability to reduce the shadow functional dimensions of the two flasks were identical effect and, thereby, improve the rate of photosynthetic (Fig 1 and Table 1). production, when compared to the standard, cylindrical Figure 1. Representation of the cross sections of the two flasks. The control flask (a) on the left, and the prototype flask (b) on the right have identical cross-sectional areas, but the sidewall configuration of the two flasks is different. These diagrams are intended for illustrative purposes only. They are not presented to scale, nor are they intended for manufacturing or engineering purposes. The key geometric measures of the respective flasks are presented in Table 1. APPLIED PHYCOLOGY 3 Table 1. Key Geometric Measures of the Flasks. included. The mean intensity applied to each culture ™ −2 −1 Geometric Measure Control AlgaTube Units was 55 µmol photons m s and was identical between Capacity Volume 1441 1441 ml the two cultures. The lights were maintained 24 hours b 2 Photon Surface Area 743 1641 cm a day with no period of darkness. Relative Photon Surface Area 100% 220% % of control d 2 Cross Section Surface Area 47 47 cm The two cultures were exposed to a continuous mag- −1 e −1 PSA Vol Ratio 0.52 1.13 cm −1 f netic field of approximately 300mT (milliTesla) each Relative PSA Vol Ratio 100% 220% % of control a 3 (Fig 2). The magnets (K&J Magnetics) were Capacity Volume: total volume of the flask when full to brim (ml = cm ). b 2 Photon Surface Area: total surface area of the sidewalls of the flask (cm ). Neodymium Grade N52 with a Br rating of 14 800 max ac Relative Photon Surface Area: expressed as %, with control = 100%. d Gauss and a BH rating of 52MGOe. Pure gaseous CO max 2 Cross Section Surface Area: area across which gas exchange and evapora- tion occur (cm ). was supplied from a pressurized tank through a regulator e −1 2 3 −1 PSA Vol Ratio: ratio of PSA to Volume for each flask (cm /cm = cm ). (FZone Model#FZ-2020PRO), a pH controller f −1 Relative PSA Vol Ratio: expressed as % with control = 100%. (Milwaukee MC122Pro), and a sparger stone (CR Brewbeer, 0.5um Diffusion Stones). The CO was dosed Culture species and growth conditions as needed using the automated pH controller set for 9.0 ± 0.2. Ambient air was continuously bubbled through both Seed culture was purchased from Algae Research Supply cultures using a HITOP Aquarium Air Pump (Haisen). in Carlsbad, CA, USA, who identify the strain only as Dissolved oxygen levels were consistently lower than Arthrospira platensis (A. platensis). Modified Schlosser 150% of ambient levels. The temperature was maintained Medium (da Silva et al., 2016) was prepared using deio- at 32°C. The Relative Humidity fluctuated mostly within nized water from the Brigham Young University (BYU) a range of 10–15%. Periodically, due to the passage of Life Science Building (LSB). Two YOHAYOH LED storm systems, the Relative Humidity fluctuated outside Light Panels (45W, 50-60 Hz, 300 mA, 110 V, 30.99 cm that range. The setup is shown in Fig 2. x 30.99 cm x 4.06 cm) were fixed to the inside of the incubator. They emitted narrow spectral peaks at 460 nm (blue) and 630 nm (red) and a substantially lower Assessment of biomass concentration level of full visible spectrum background radiation. The ratio of red to blue in the present study is calculated Prior to daily sampling, each culture was mixed manu- from the manufacturer’s data as approximately 60:40. ally to promote homogeneity. Each day, the cultures A 15-Watt incandescent bulb (General Electric) was also were tested for Dissolved Oxygen (RCYAGO Portable Figure 2. Configuration of the growth chamber. The control flask was on the left, and the prototype flask was on the right. The two light panels were affixed to the rear wall of the incubator. The incandescent light was attached to the door and centered. The magnets can be seen on the lower left side of the control flask and on the lower right side of the prototype flask. The picture was taken with the door open for convenience. 4 J. L. GAL ET AL. Table 2. Dilutions for % Transmittance vs. Concentration stan- Dissolved Oxygen, Model#DO9100), pH (Milwaukee dard curve. MC122Pro), Total Dissolved Solids and Electrical –1 Dilution # Dilution Rate (%) Concentration (g l ) %T ABS Conductivity (Vivosun TDS/EC Metre), and Light Stock 100 1.63 0.70 >2.02 Transmittance and Absorption (Biolog Turbidimeter, 1 61 1.00 0.90 >2.02 −1 Model#21907, 590 nm). Concentration (C g l ) was 2 55 0.90 0.95 2.02 3 49 0.80 1.00 2.00 calculated from %T using a standard curve. 4 43 0.70 1.20 1.95 The organisms were observed to form clumps of 5 33 0.60 1.99 1.80 6 31 0.50 3.10 1.52 filaments, a sign of their previously documented colo- 7 25 0.40 5.70 1.30 nial nature (Demoulin et al., 2019). The clumps were 8 18 0.30 10.50 0.98 9 12 0.20 23.50 0.64 a source of minor error for the OD readings because 10 6 0.10 47.50 0.33 they interfered with homogeneity. The clumps caused 11 3 0.05 69.50 0.16 12 1 0.02 85.50 0.07 some minor, but noticeable, drift in the turbidity read- ings, as they circulated in the culture samples. To reduce the margin of error associated with this imperfect The diluent for these dilutions was Zarrouk Media homogeneity, the turbidity metre was observed for (ZM), because ZM was being used at the time the a full minute for each reading. Time weighted means standard curve was prepared. Subsequent to the creation of the %T and ABS were recorded for each mea- 590 590 of this standard curve, a decision was later made to surement, rather than single spot readings. switch to MSM media. To examine the potential impact that the change in diluent might have on the OD read- ings, the OD for pure MSM was compared with the Standard curve preparation OD for pure ZM. It was determined that the differ - A stock of A. platensis ARS was taken from a well-mixed ence in %T readings between the two media (<2%) was culture that had previously plateaued in the incubator. sufficiently small to justify continued use of the standard Five samples of 10 ml each were pipetted from that curve, without preparation of a new standard curve stock, and each was measured using the dry weight using MSM as the diluent. protocol (supplementary material). Those five samples Table 2. The data for the standard curve presented in produced five different results for dry weight − 1.63 g Fig 3. These data establish a correlation between bio- −1 −1 −1 −1 −1 −1 l , 1.68 g l , 1.34 g l , 1.71 g l , and 1.57 g l – which mass concentration (g l ) of A. platensis and the optical reflected various sources of error in the assay (supple- density of the culture at 590 nm. “%T” stands for mentary material). The high and low values from the Percentage Light Transmission. “ABS” stands for range were discarded, and the mean of the remaining Absorbance in absorbance units. %T and ABS are inter- −1 three values was calculated to be 1.63 g l . The 95% related, according to the standard Beer-Lambert equa- confidence interval of the three sample dry weights was tion: ABS = 2 – log (%T × 100). calculated to be ±3.83%. This was determined to be sufficiently accurate for the present study. Calculation of production (P ) Another sample from the same stock was taken and measured in the turbidimeter, producing a %T reading P is calculated by subtracting the previous day’s C from of approximately 0.70%. From this sample, twelve dilu- the current day’s C as follows. −1 tions were made to produce values of 1.00 g l down to −1 P dðg per lÞ ¼ C dðg per lÞ C d 1ðg per lÞ (1) 0.02 g l , and each dilution was tested in the turbidi- meter for %T with mass being calculated based on the P can be calculated for a multi-day time-period, in −1 dilution, using the 1.63 g l figure as a reference point. which case it is termed “Cumulative Production” (CP −1 These data are show in Table 2. A standard curve was in g l ), and it can also be done for each single day, in made from the data (qryyig 3). The table was then used which case the value is termed Daily Production (DP in −1 to convert each day’s %T readings into a corresponding g l ). In the time parsed data analysis presented in the C data point for each flask. For samples with particu- results section, DP values are calculated for each day −1 larly high C (generally anything above 0.70 g l ), sam- and then averaged together longitudinally for particular ples were diluted prior to determination of C, because % time periods. Those means are termed Mean Daily −1 T readings were more difficult to discern at levels below Production (MDP in g l ) for the time-period in 1%. Generally, the dilution rate was 1 : 5. Such samples question. were assessed at 1 : 5 dilution and that result was then As a metric, P (and its associated metrics: CP, DP, multiplied by 5x to reach the final value. and ADP) focus on the absolute amount of biomass APPLIED PHYCOLOGY 5 0.02, 85.5% 100% 0.05, 69.5% 0.10, 47.5% 0.20, 23.5% 0.30, 10.5% 10% 0.40, 5.7% 0.50, 3.1% 0.60, 2.0% 0.70, 1.2% 1% 0.80, 1.0% 0.90, 1.0% 1.00, 0.9% 0% 0.00 0.25 0.50 0.75 1.00 -1 Biomass Concentra!on (g l ) Figure 3. Standard Curve. This chart plots the data from the dilution table to illustrate the standard curve used to determine daily culture concentration from OD readings. produced for a particular time period, regardless of the compounding effect and is presented as a compound initial concentration of the culture. Other metrics, such mean rate over multi-day time periods. The mathema- as Specific Growth Rate (SGR), focus on the percentage tical equation used to calculate SGR of biomass for of biomass growth per day, which depends on the initial a particular time-period (d to d ) is presented, here. 0 10 � � C . The difference between the two metrics is best C10 C10 Ln Ln C0 C0 observed by examining their respective units. SGR d0 d10 ¼ ¼ (2) d10 d0 10 SGR is presented as a fractional value per day (e.g., −1 0.20 day ). This would mean that the culture grew at Mathematically, SGR can be calculated for any time- a compound mean rate of 20% per day during the period in an experiment, but the common practice in measured time-period. On the other hand, P is pre- phycology is to present SGR for the entire length of the −1 −1 sented in g l day . Biomass is sold and consumed by experiment and to also include the presentation of weight, not by percentages or by fractions. Therefore, P SGR . SGR is the maximum of the daily SGR d max max may be more the more important metric for industry. results for each culture. It is an indicator of the best On the other hand, SGR is widely published in scien- single day’s performance. tific journals, and academic readers may prefer this The abbreviation we used for Specific Growth Rate is metric. When it is presented as a fraction of the SGR, which is different from the abbreviation used in Concentration, and for multi-day periods, then it takes other studies (μ). The reason for breaking from conven- the effect of daily compounding into account. Since dif- tion is that the Greek letter “μ” is used in this study to −2 ferent readers may have different preferences on how best describe light intensity in the units of μmol photons m −1 to measure biomass growth, both metrics are presented s . To avoid confusion in the present study, “μ” is herein, and readers are invited to decide for themselves restricted only to units of light intensity, and Specific which metric to use for their own purpose(s). Growth Rate is abbreviated as SGR. Calculation of specific growth rate (SGR) Calculation of Delta (Δ) SGR is based on the mathematical concept of the natural To measure performance differences between the two logarithm (Ln). The amount produced per day is pre- vessels (control and prototype), Δ was calculated using sented as a fraction of the total concentration on the the control datasets and the prototype datasets. Δ is previous day, rather than as an absolute value. For expressed in percentage terms, relative to the control. multi-day time periods, it takes into account the It is not specific to any particular metric. Δ can be % Light Transmi"ance (590nm) 6 J. L. GAL ET AL. −1 −1 calculated for any metric and any time-period and (MxDP) in the prototype, was 0.29 g l day , 38% labelled accordingly. For example, the difference greater than the comparable figure for the control, −1 −1 between the prototype culture concentration and the 0.21 g l day . The MxDP results were both achieved −1 control culture concentration (C in g l ) on day #5 on different days than SGR , which highlights the max would be labelled Δ . The difference in cumulative difference between the two metrics. While theoretical C5 −1 production (CP in g l ) on day #8 would be labelled phycologists may be interested in the greatest daily Δ . And the difference in Mean Daily Production percentage growth, industrial producers may be more CP8 −1 −1 (MDP in g l day ) during the first seven days would focused on the greatest daily mass growth, because they be labelled Δ . The equation for Δ of Cumulative MDP1-7 Production on day #d (Δ ) is presented here. CPd ΔCPdð%Þ100%� ½CP pdðg per lÞCP cdðg per lÞ�� ½CP cdðg per lÞ� (3) It should be noted, here, that lower-case “c” and lower- case “p” denote control and prototype, respectively. They should not be confused with upper-case “c” and upper-case “p” which denote Concentration and Production, respectively. Calculation of statistical significance (p-values) The datasets were determined to follow non-normal distributions, which prevented the use of the standard t-test. Therefore, a non-parametric test was used. Statistical significance was assessed using the Wilcoxon Rank Sum Test (Mann-Whitney U Test; Corder, Foreman, Wiley, Sons, & Hoboken, 2014). The resulting p-value was considered statistically significant at p < 0.05. Results Both the control and prototype concentration data exhibited non-linear trends, and the two flasks each produced different shaped growth trends. A quadratic model was fit to the control data. A non-constant var- iance linear spline regression model was fit to the pro- totype data. The prototype data points from the four independent replicates showed a gradually increasing variance as the day number increased, a result that was not observed to a similar extent in the control flask. The means, however, still showed statistically significant Figure 4. (A) and 4(B). Daily Concentration and Cumulative growth differences in favour of the prototype. (Fig 4). Production values of prototype and control tubes. 4(a) Chart Specific Growth Rate (SGR) was calculated as −1 −1 plotting the daily Concentration (C ), averaged across the four 0.17 day for the control flask and 0.20 day for the independent replicates, for the AlgaTube™ prototype and prototype flask, a difference of 17% in favour of the cylindrical control, respectively. The prototype data points are AlgaTube™. SGR was also calculated for both flasks, max represented by solid lines and black triangles (━▲━). Δ = C10 −1 −1 and the results were 0.40 day and 0.49 day for the +34% (p = 0.02, Wilcoxon Rank Sum Test). 4(b) Chart plotting the daily Cumulative Production (CP ), averaged across the four control and prototype, respectively. Both SGR data- d max independent replicates, for the two vessels, AlgaTube™ proto- points occurred on Day #1, when the cultures were at type and cylindrical control, respectively. δ = +41% (p = cp10 their lowest concentrations. Daily Production (DP ) was 0.02, Wilcoxon Rank Sum Test). In both 4(a) and 4(b), the calculated, and the resulting dataset focuses attention on prototype data points are represented by solid lines and black the amount of biomass produced per day, rather than triangles (━▲━). The control data points are represented by dashed lines and black circles (—□—). the cumulative total. The Maximum Daily Production APPLIED PHYCOLOGY 7 sell product by weight. These two metrics are different, Discussion and they do not occur on the same day. Growth conditions To compare the Mean Daily Production (MDP) for different time periods, the Mean Daily Production data- The purpose of this experiment was to isolate and set was parsed into three separate time periods: (1) the evaluate the shape of the sidewalls of the AlgaTube full 10 days, (2) the first seven days, and (3) the last three prototype flask for its effect on growth rates. This days. A longitudinal mean was calculated for each time required all other variables to be held constant and period. Between the first seven days and the last three identical between the control flask and the prototype days, Mean Daily Production (MDP) in the control flask flask. It also required that no other variable be set to −1 −1 −1 dropped by 38% from 0.13 g l day down to 0.08 g l a level that was rate limiting or restrictive of the −1 day ; and this caused a large increase in the Delta (Δ) growth of the organism. The existence of any single during the final three days. At +113%, Δ was rate limiting factor, other than the PSA of the respec- MDP8-10 nearly five times higher than Δ . (Fig 5). tive culture flasks, could inhibit both cultures growth MDP1-7 The time-parsed data analysis clearly shows that the rates to a degree that it would override and conceal the biomass growth in the control flask decreased in the hypothesized positive effect of the enhanced surface final three (3) days of the experiment, whereas there area. Thus, to evaluate the singular effect of the shape was no such decrease in the prototype flask. This was of the sidewalls, the value or setting of the other vari- most likely due to a negative impact caused by the ables was a matter of careful consideration and subject shadow effect. The filaments shaded each other when to pre-testing via preliminary experiments. their concentrations increased. On the other hand, the Other researchers have shown large variations in prototype did not exhibit the same negative impact growth curves and production rates of A. platensis by during that same time-period (Fig 5). When viewed using different ratios of red and blue light, while holding −2 −1 alongside the Delta (∆) values noted in Fig 4, this total intensity constant at 100 μmol photons m s . observation suggests that the enhanced surface area of Maximum growth was achieved with a red-to-blue ratio the prototype reduced the shadow effect and enabled the of 70: 30. (Lima, Teixeira, Teixeira, Filócomo, & Lage, culture to maintain a higher rate of growth and for 2018). The ratio of red to blue in the present study is a longer time-period, when compared to the cylindrical calculated from the manufacturer’s data as approxi- control flask. mately 60:40. To enhance stimulation of P700, Figure 5. Time-Parsed Mean Daily Production. The Mean Daily Production (MDP) data were, themselves, averaged together long- itudinally, for each of three different time periods within the 10-day experiment. The respective Delta (∆ ) values and p-values for mdp different time periods are shown. The mean culture concentration during each of those time periods is shown below the x axis labels. The bars representing the control flask are white, and the bars representing the prototype flask are shaded grey. 8 J. L. GAL ET AL. a photoreceptor that absorbs 700 nm light, a 15-Watt However, at 55% relative humidity, the growth rates of incandescent bulb (General Electric) was added the cultures were also significantly reduced, compared to (Webber & Lubitz, 2001). Incandescent bulbs produce the growth rates observed at 10% relative humidity, with- a broad spectrum of radiation that is heavily weighted out the water tubs. This was attributed to the light towards the red, far red, and infra-red regions of the attenuating effect of humidity, the same effect observed visible spectrum (Azizi, Golmohammadi, & Aliabadi, in the solar insolation data of the National Solar −1 2016). All of the light sources were on 24 hr day , Radiation Database of the NREL. To avoid the inhibitory according to the method of Prates, Radmann, Duarte, effect of high humidity in the present study, it was deter- de Morais, & Costa (2018). mined not to use the tubs of water. In lieu of tubs to Using an enhanced silicon photodiode assembly (LI- reduce evaporative water loss, deionized water was added 190 R Quantum Sensor), the light intensity was mea- daily to replace evaporative water loss in the cultures. sured at 24 spot locations around the perimeter of the Kazbar et al. (2019) showed that, when dissolved flasks, or 12 locations per flask. Spot readings ranged oxygen (DO) levels reach approximately 300% of atmo- −2 −1 from 22–105 µmol photons m s . This wide range of spheric concentration, significant inhibition of the light intensity was the natural outcome of the physical growth rate of photosynthetic cultures of Chlorella vul- placement of the light sources and the culture flasks garis was observed. Torzillo & Vonshak (2013) within the incubator. The sides of the culture flasks described a cascade of inhibiting effects triggered by that faced directly towards the light sources experienced excess O , and its effect on the concentration of a higher intensity than the sides not facing the light Reactive Oxygen Species (ROS) such as superoxide radi- sources directly. This kind of intensity imbalance is cal (O –), hydrogen peroxide (H O ), and hydroxyl 2 2 2 consistent with what is found in outdoor tubular photo- radical (·OH). Ganesh, Manoharan, & Suraishkumar bioreactors (tPBRs) that are illuminated by the sun. (2007) showed that large increases in ROS correlated Li, Guo, Li, & Cai (2007) found that an electromag- strongly with reductions in biomass growth rates in netic field enhanced the growth rate of A. platensis at an Spirulina maxima. −2 −1 intermediate light intensity of 252 μmol photons m s . Using the cyanobacterium Synechocystis sp. PCC Deamici, Costa, and Santos (2016) also reported positive 6803, Nishiyama, Allakhverdiev, Yamamoto, Hayashi, effects on A. platensis growth rates from magnetic fields, & Murata (2004) demonstrated that another ROS, sing- as did de Costa Menestrino et al. (2021). Based on those let oxygen ( O ), appears to inhibit the repair of photo- two studies, it was determined to use magnets in this damaged PSII by inhibiting production of a key experiment. The temperature was derived from component of PSII – the D1 protein (Aro, Ivar, & a previously published range of preferred temperature Andersson, 1993) - at the translational level. This feed- for A. platensis (Oliveira, Monteiro, Robbs, & Leite, back loop has the effect of shutting down the function- 1999; Soni, Sudhakar, & Rana, 2019). ality of PSII, which is to split water molecules into As we observed during the preliminary studies that hydrogen atoms and O (Aro, Ivar, & Andersson, preceded these trials, humidity can have two effects on 1993). This being the first step of the photosynthetic cultures. First, it impacts the rate of evaporation of pathway, the rate of photosynthesis is reduced by these water from the cultures (data not included). Second, events. These biochemical events also reduce the pro- humidity can interfere with the transmission of light duction of O by PSII, thus countering the excess O 2 2 to the cultures. This latter effect can also be seen in the levels that caused them in the first place. This is a self- natural world by examination of the solar insolation fulfiling and self-protective feedback loop. data in different parts of the world, where significant Another impact of excess oxygen levels in photosyn- variation in annual solar insolation data at ground level thetic microorganisms is the stimulation of a secondary is found to correlate inversely with the mean humidity metabolic pathway, photorespiration, in which the in different areas at the same latitude (National Solar Rubisco enzyme utilizes oxygen in lieu of carbon Radiation Database, National Renewable Energy (Vonshak, Torzillo, Accolla, & Tomaselli, 1996; Fernie Laboratory, U.S. Government). & Bauwe, 2020; Sforza et al., 2020). There is some The light attenuating effect of humidity can also be evidence that the O : CO ratio, more-so than the 2 2 observed in small, experimental systems in a laboratory. absolute level of dissolved oxygen, governs this pathway During pre-testing for this experiment, tubs of water (Kitaya, Azuma, & Kiyota, 2005). Nevertheless, the net were included to reduce evaporation inside the incubator. effect of excess oxygen can be summarized as With two large tubs of water in the incubator, relative a combination of shutting down photosynthesis and humidity was maintained at approximately 55% and turning on photorespiration. Based on observations of evaporation from the cultures was greatly reduced. reduced biomass production rates at high dissolved APPLIED PHYCOLOGY 9 oxygen levels (Kazbar et al., 2019 and preliminary test- the expression rate of the phycobiliproteins between the ing for the present study), the photorespiratory pathway control and the prototype. However, the standard curve was assumed to be much slower, kinetically, than covered a broad range of culture concentrations from −1 −1 photosynthesis. 0.02 g l up to 1.0 g l . Therefore, the standard curve is During pre-testing for the present study, cultures believed to have fully accounted for any differences that were also observed without supplemental CO . The pH may have been caused by differences in concentration. of those cultures drifted steadily higher from a starting In terms of light intensity, Nomsawai, de Marsac, level of 9.0 up to a level of 11.0, or higher in some cases. Thomas, Tanticharoen, & Cheevadhanarak (1999) did A concomitant plateauing of daily growth was observed show significant variability in phycobiliprotein expres- as the pH increased. These observations were consistent sion in A. platensis C1 when the light intensity was with a report of the optimal pH for A. platensis total changed by a large factor of 10x from 50 μmol −2 −1 −2 −1 biomass growth being pH 9.0 (Ismaiel, El-Ayouty, & photons m s to 500 μmol photons m s . Piercey-Normore, 2016). However, Rizzo et al. (2015) showed that smaller changes in light intensity from 50 to 100 to 150 μmol −2 −1 photons m s did not have a statistically significant OD and phycobiliprotein expression 590 −1 effect on the level of total protein content (%mg mg , p −1 The use of OD to measure optical density of cultures < 0.05) or on the phycobiliprotein content (%mg mg , in this study raises the possibility of errors in the mea- p < 0.05) in cultures of A. platensis. The present study surement of C , because 590 nm is near the spectral was conducted at low light intensity, and well below 150 −2 −1 absorbance peak of the phycobiliproteins, an important μmol photons m s . Thus, differences in phycobili- class of photopigments expressed by A. platensis (Barber protein expression most likely did not impact the C & Richards, 1977; Kronick, 1986). Differential expres- results. sion of these photopigments between the control culture and the prototype cultures could have been, theoreti- Phycological consistency of results cally, a source of error in the OD readings, because changes in OD could be attributed to differences in The higher rate of biomass production observed in the photopigment expression, rather than differences in prototype compared to the control is conceptually con- total biomass concentration. However, this possible sistent with results reported by other researchers in the −1 source of error can most likely be ruled out by the field of phycology. The application of a higher PSA vol following methods and logic. ratio caused improvement in both C and P of d d In the present study, the control and prototype cul- A. platensis UTEX 1926, according to da Silva et al. tures were established from a common culture, the (2016). Converti, Lodi, Del Borghi, & Solisio (2006) “starter” culture. Prior to division of the starter culture showed that cultures of A. platensis UTEX 1926 grew into the control flask and the prototype flask, it was faster and to higher plateaus in tPBRs with greater PSA −1 mixed thoroughly to ensure uniformity between the vol ratios than in an open pond. The same phenom- control and prototype cultures on Day #0. Any differ - enon was also observed in Chlorella vulgaris CCAP211 by ences in the rate of expression of the phycobiliproteins, Frumento et al. (2013). In all of these studies, including −1 therefore, would have to have emerged during the 10- the present study, a higher PSA vol ratio helped to day trial period, not prior to it. Since the cultures were reduce the self-shading effect and to expose greater num- identical on Day #0, the only possible causes for differ - bers of cells, filaments, and/or photoreceptor pigment ential expression of the phycobiliproteins during the 10- molecules to incoming photons. In our time parsed day trials were: (1) differences in C as the cultures grew data (qryyig. 5), we elaborate on the previously known at different rates and (2) differences in the light intensity self-shading effect by showing that it actually has two caused by the PSA difference between the two flasks. All components: (1) a slower rate of growth during the first of the other variables, and inputs, without exception, 7 days and (2) avoidance of the substantial growth rate were fixed and controlled so that they were equal reduction seen in the control flask during days #8–#10. between the two flasks. In all of these studies, compelling evidence and com- As the growth data show, the C in the prototype parisons are presented, and we can learn more about the flask was consistently higher than the C in the control performance of the AlgaTube™ by comparing the various flask. Therefore, it is theoretically possible that the dif- studies to each other and to our own data. It is note- ference in concentrations between the two cultures worthy, for example, that da Silva et al. (2016) showed −1 could have triggered an internal, biochemical pathway a C of 8.44 g l after only 9 days of growth. At max in the organism, which, in turn, caused a difference in +344%, this C was far higher than the present study, max 10 J. L. GAL ET AL. −1 which showed a C of 1.90 g l after 10 full days of remaining difference, +130%, can be attributed to the max growth. There were several important differences in con- combined effect of the other remaining variables, a list figuration, which may explain the difference. that includes: light quality (fluorescent vs. LED), vessel The da Silva study used a higher starting concentra- material (glass vs. 3-D printing resin), supplemental −1 −1 tion on Day #0 of 0.40 g l (60% higher than 0.25 g l hardware (the hose clamps on the AlgaTube™ blocked for the present study). It was run with a higher light 7% of the PSA), and mixing technique. −2 −1 intensity of 100 μmol photons m s (82% higher than Some of these variables can be approximated using −2 −1 55 μmol photons m s for the present study). The known information. Geometrically, the hose clamps −1 −1 reported PSA vol ratio was 1.94 cm (70% higher in block about 7% of the PSA; so, that effect is estimated −1 than 1.13 cm for the present study). There were also at 7%. The difference in the light transmission of glass differences in the quality of the light (fluorescent vs. vs. the 3-D printing resin is known to be about 10% LED), the vessel materials (glass vs. 3-D printing resin), (with some variation depending on wavelength); so, and the supplemental hardware (the hose clamps on the that effect is estimated at 10%. Based on analysis of AlgaTube™ blocked 7% of the PSA). Finally, and perhaps two different studies (Chainapong et al., 2012; most importantly, there was a very important difference Thaweedet et al. 2012; Lima, Teixeira, Teixeira, in the mixing techniques between these aqueous cultures. Filócomo, & Lage, 2018), light quality is estimated to To whit, by circulating their cultures continuously have an impact of 30%. That leaves approximately through a looped pipe network, da Silva et al. (2016) 83% as the remainder, which can be attributed to used a different mixing technique and introduced another the mixing technique. This analysis concludes, then, variable to their comparisons. They compared a horizontal that the mixing technique is the single most important tPBR with a circulating loop configuration against a simple difference between the two studies, though the next shaken flask and a paddled raceway. Not only were the three variables are not far behind. The final determi- −1 PSA vol ratios different between those three configura - nation of the relative effect of all of the differences tions, but there were also three different mixing techniques between the da Silva study and the present study are used: continuous circulation, flask shaker machine, and presented and summarized in Table 4. paddle wheel. Their conclusions, therefore, should be NB: Strain differences were not considered in this understood in light of these important differences in mix- table or in the related discussion. ing technique and not solely as a reflection of the different −1 PSA vol ratios. Indeed, the different mixing techniques Light containment in that study raise an important question about which variable was most important to their exceptionally high Light containment is not considered as a separate vari- −1 growth rate: the increased PSA vol ratio, the continuous able unto itself. Rather, it is a means to increase the light circulation, or the combination of the two. intensity without increasing the power input to the In the present study, there was no continuous circula- lights. Since light intensity is already included in the tion in either culture. The only mixing applied to both evaluation of the key variables above in Section 4.2, cultures in the present study was provided by the energy our discussion of light containment is presented here of the air bubblers, and that was identical in both cultures. separately, not as part of the list of variables in the Thus, the present study was more precisely controlled for previous section. −1 PSA vol ratio as the sole variable in the experiment. The The light intensity inside the incubator of this −1 important question of the relative effects of PSA vol vs. study was considerably different with the door open, mixing technique arising out of da Silva et al. (2016) remains unanswered, empirically. However, it is possible Table 3. Quantification of primary differences between present to derive an estimated answer to that question, mathe- study vs. da Silva et al. (2016). matically, based on data that are currently available. Value in Value in Present da Silva et al. Delta First, the differences in configuration between the da Variable Name Study (2016) (Δ) Silva study and the present study are quantified and −1 Initial Concentration [g l ] 0.25 0.40 +60% added together to reach an estimate of their cumulative Light Intensity [μmol 55 100 +82% −2 −1 photons m s ] total effect on C as shown in Table 3. max −1 −1 PSA Vol Ratio [cm ] 1.13 1.94 +72% Assuming that the effect of these differences on the Total Delta (Δ) +214% growth metrics are additive (they may not actually be), Maximum Concentration [g 1.90 8.44 +344% −1 l ] the three primary variables listed in Table 3 can be esti- In the present study, C was on Day #10. In da Silva et al. (2016), C was max max mated to have accounted for a total of +214% out of the on Day #9. ab +344% difference in C between the two studies. The A positive Δ means da Silva et al. (2016) was higher than the present study. max APPLIED PHYCOLOGY 11 Table 4. Ranked summary of all differences between present may be dependent, however, on the angle of incidence of study vs. da Silva et al. (2016). the solar radiation on the light containment housing, an Value in Value in effect that has been very thoroughly studied in the field of Present da Silva et al. Delta photovoltaics (PV). Solar panels are well known to per- Variable Name Study (2016) (fjfj) Mixing Technique Air Bubbles Continuous +83% form best when angled continuously towards the sun Circulation with a device that rotates throughout the day to follow Light Intensity (µmol 55 100 +82% the sun, or when set at a fixed angle called “Latitude Tilt” photons m-2 s-1) −1 −1 PSA Vol Ratio (cm ) 1.13 1.94 +72% which maximizes the intensity of the sun by averaging −1 Initial Concentration (g l ) 0.25 0.40 +60% out its different seasonal phases and daily paths (Nicolás- Light Quality LED Mix Cool White +30% Fluorescent Martín, Santos-Martín, Chinchilla-Sánchez, & Lemon, Vessel Material 3-D Printing Glass +10% 2020). It is expected that outdoor PBRs with light con- Resin Extra Hardware Hose Clamps n/a +7% tainment housings may behave analogously. Indeed, this Total Delta +344% may be one of the key reasons that outdoor systems are a −1 C (g l ) 1.90 8.44 +344% max a widely known to underperform lab studies. In the present study, C was on Day #10. In da Silva et al. (2016), C was max max on Day #9. ab A positive Δ means values in da Silva et al. (2016) was higher than the present study. Summary of key metrics This study demonstrates that the AlgaTube™ clearly compared to the door being closed. With the door outperformed the control flask in all the key metrics open, the iPPFD was measured to be 29 µmol used for comparison. In Table 5, the Delta (Δ) values are avg −2 −1 photons m s , or 55% lower than the iPPFD of presented relative to the control, meaning that positive avg −2 −1 55 µmol photons m s when the door was closed. Delta (Δ) values show that the AlgaTube™ performed This observation is noteworthy for the field of PBR better than the control. Across the entire spectrum of design, because it shows that light containment has growth metrics, the AlgaTube™ consistently outper- a large impact on the mean radiation received by formed the control flask (Table 5). phycological cultures in artificial systems. Without Finally, the decrease in the biomass growth in the containment, a large percentage of PAR photons can control flask during the final three days of the experi- be lost to the surrounding environment without ever ment, which was not seen in the prototype flask, sup- being used for biomass growth. The horizontal tPBR ports the hypothesis that the prototype flask’s gains are of Converti, Lodi, Del Borghi, & Solisio (2006), for due to overcoming the shadow effect. In the control example, did not include any light containment fea- flask, the filaments shaded each other, when their con- tures and might be improved upon by including centrations increased. On the other hand, the prototype a housing made of white walls which reflects and did not exhibit the same negative impact during that contains stray photons. same time-period (Fig 5). When viewed alongside the This concept of light containment has been success- Delta (∆) values noted in Fig 4, this observation suggests fully used for land plants for many years (Warman & that the enhanced surface area of the prototype reduced Mayhew, 1979); and therefore, it is likely to have similar, the shadow effect and enabled the culture to maintain positive effects on aqueous photosynthetic cultures. At a higher rate of growth and for a longer time-period, present, however, state-of-the-art, outdoor PBRs gener- when compared to the cylindrical control flask. ally do not include any light containment features. The comparative analysis of different time peri- It is reasonable to hypothesize that the inclusion of ods showed also that the prototype improved the light containment features on outdoor systems may Average Daily Production by 113% during the final increase the utilization of physically limited solar radia- three days of the trial, when the cultures were at tion by, in effect, “recycling” stray photons. This effect their thickest. Table 5. Summary of key metrics. Key Metric Control AlgaTube™ Delta (Δ) p-value −1 Maximum Concentration (g l ) 1.42 1.90 +34% 0.02 −1 Total Cumulative Production (g l ) 1.17 1.65 +41% 0.02 −1 Specific Growth Rate (day ) 0.17 0.20 +17% 0.02 −10 Maximum Specific Growth Rate (day ) 0.40 0.49 +23% 0.02 −1 Mean Daily Production (g l ) 0.12 0.17 +41% 0.001 −1 Maximum Daily Production (g l ) 0.21 0.29 +38% 0.19 a TM Positive values for Delta indicate that the AlgaTube was higher than the control. 12 J. L. GAL ET AL. Future directions laboratory experiments were performed by JLG, who also wrote the first draft of the manuscript. NRC and DLE per- The present study serves as compelling proof-of- formed the statistical analyses, created the charts, and wrote concept for the novel shaped walls of the AlgaTube™. the statistical language. Subsequent drafts were reviewed and edited by all four authors and by both reviewers and an editor The prototype performed better than the control on all at the Journal of Applied Phychology. six growth metrics as summarized in Table 5. This study is considered a successful first step, but there remains considerable potential to optimize the performance of References such tubes, as shown by the comparisons to other stu- dies summarized in Table 4. Adeniyi, O. M., Azimov, U., & Burluka, A. (2018). Algae The AlgaTube™ should be tested, for example, in Biofuel: Current status and future applications. Renewable and Sustainable Energy Reviews, 90, 316–335. doi:https:// a circulating loop, horizontal configuration such as the doi.org/10.1016/j.rser.2018.03.067 tPBR configuration demonstrated by da Silva et al. Aro, E.-M., Ivar, V., & Andersson, B. (1993). Photoinhibition (2016). It should be tested in smaller diameter sizes, of photosystem ii: Inactivation, protein damage and turn- such as those used by Converti, Lodi, Del Borghi, & over. Biochimica Et Biophysica Acta (BBA) – Bioenergetics, Solisio (2006), and in a circulating loop vertical config - 1143, 113–134. doi:https://doi.org/10.1016/0005-2728(93) uration. The important factor of light containment 90134-2 Arora, K., Kumar, P., Bose, D., Li, X., & Kulshrestha, S. (2021). should be studied further. Vertically oriented tubes con- Potential applications of algae in biochemical and bioe- tained in a containment housing, for example, might nergy sector. 3 Biotech, 11, 1–24. doi:10.1007/s13205-021- perform better than stand-alone vertical tubes in a fully 02825-5 transparent greenhouse. Continuous feeding and fed- Azizi, M., Golmohammadi, R., & Aliabadi, M. (2016). batch methodologies should also be tested in the Comparative analysis of lighting characteristics and ultraviolet emissions from commercial compact fluorescent and incan- AlgaTube™. And, finally, as with any new PBR concept, descent lamps. Journal of Research in the Health Sciences, 16, the transition to larger scale, outdoor environments 200–205. https://pubmed.ncbi.nlm.nih.gov/28087852 must also be made. Barber, D. J. W., & Richards, J. T. (1977). Energy Transfer in the Accessory Pigments R-Phycoerythrin and C- Phycocyanin. Photochemistry and Photobiology, 25, 565– Acknowledgements 569. doi:10.1111/j.1751-1097.1977.tb09129.x Benemann, J. R. (1979). Production of nitrogen fertilizer with We thank Brigham Young University for providing nitrogen-fixing blue - green algae. Enzyme and Microbial laboratory space, laboratory equipment, and general Technology, 1, 83–90. doi:https://doi.org/10.1016/0141- laboratory supplies. We thank the first author, JLG, for 0229(79)90103-0 his personal contribution of financial support, which was Chainapong, T., Traichaiyaporn, S., & Deming, R. L. (2012). used for prototyping manufacturing expenses and the Effect of light quality on biomass and pigment production purchase of certain laboratory instruments. We thank in photoautotrophic and mixotrophic cultures of Spirulina Brigham Young University’s Engineering Department platensis. Journal of Agricultural Technology, 8, 1593–1604. for use of its Prototyping Lab. We thank Mr. Nicolas Chong, J. W. R., Khoo, K. S., Yew, G. Y., Leong, W. H., Miller, draftsman, who was hired by JLG to make the Lim, J. W., Lam, M. K. . . . Show, P. L. (2021). Advances in technical drawings of the prototype and Mr. Harlan production of bioplastics by microalgae using food waste Stevens, BYU undergraduate, who proofread the manu- hydrolysate and wastewater: A review. Bioresource script without pay. Xometry™ and Stratasys™ were hired Technology, 342, 125947. doi:10.1016/j.biortech.2021.125947 by JLG for 3-D printing of the vessels. Christaki, E., Karatzia, M., & Florou-Paneri, P. (2010). The use of algae in animal nutrition. Journal of the Hellenic Veterinary Medical Society, 61, 267–276. doi:10.12681/ Disclosure statement jhvms.14894 Clippinger, J., & Davis, R. (2019). Techno-Economic analysis No potential conflict of interest was reported by the author(s). for the production of algal biomass via closed photobior- Mr. Jonathan L. Gal, Dr. Steven M. Johnson, and the Brigham eactors: future cost potential evaluated across a range of Young University Office of Technology Transfer each have a cultivation system designs. National Renewable Energy financial interest in the patent application(s) related to the Laboratory, Golden, CO. NREL/TP-5100–72716. https:// present study. www.nrel.gov/docs/fy19osti/7271 . Converti, A., Lodi, A., Del Borghi, A., & Solisio, C. (2006). Cultivation of spirulina platensis in a combined airlift- Author contributions tubular reactor system. Biochemical Engineering Journal, 32, 13–18. doi:https://doi.org/10.1016/j.bej.2006.08.013 The idea for the AlgaTube™ was conceived by JLG, individu- Corder, G. W., & Foreman, D. I. (2014). Nonparametric ally. The experiment was designed collaboratively by JLG and Statistics: A Step-by-Step Approach (Vol. 4, 2nd ed., pp. SMJ. Laboratory space, certain laboratory equipment, and 69–75). Hoboken, NJ, USA: John Wiley & Sons, Inc. general laboratory supplies were provided by SMJ. All APPLIED PHYCOLOGY 13 da Costa Menestrino, B., Sala, L., Costa, J. A. V., Buffon, J. protein amount on Chlorella vulgaris. Fresenius G., & Santos, L. O. (2021). Magnetic Fields Exhibit a Environmental Bulletin, 26, 7974–7980. Positive Impact on Lipid and Biomass Yield During Kitaya, Y., Azuma, H., & Kiyota, M. (2005). Effects of Phototrophic Cultivation of Spirulina Sp. Bioprocess Temperature, CO /O concentrations and light intensity 2 2 and Biosystems Engineering, 44, 2087–97. doi:10.1007/ on cellular multiplication of microalgae, Euglena gracilis. s00449-021-02585-9 Advances in Space Research, 35, 1584–1588. doi:https://doi. da Silva, M. F., Casazza, A. A., Ferrari, P. F., Perego, P., org/10.1016/j.asr.2005.03.039 Bezerra, R. P., Converti, A., & Porto, A. L. F. (2016). Kronick, M. N. (1986). The Use of Phycobiliproteins as A new bioenergetic and thermodynamic approach to Fluorescent Labels in Immunoassay. Journal of immunolo- batch photoautotrophic growth of Arthrospira (Spirulina) gical methods, 92, 1–13. doi:10.1016/0022-1759(86)90496-5 platensis in different photobioreactors and under different Kumar, K., Dasgupta, C. N., Nayak, B., Lindblad, P., & Das, D. light conditions. Bioresources Technology, 207, 220–228. (2011). Development of suitable photobioreactors for CO Deamici, K. M., Costa, J. A. V., & Santos, L. O. (2016). sequestration addressing global warming using green algae Magnetic fields as triggers of microalga growth: and cyanobacteria. Bioresource Technology, 102, 4945–4953. Evaluation of its effect on Spirulina sp. Bioresources doi:https://doi.org/10.1016/j.biortech.2011.01.054 Technology, 220, 62–67. doi:10.1016/j.biortech.2016.08.038 Li, Z.-Y., Guo, S.-Y., Li, L., & Cai, M.-Y. (2007). Effects of Demoulin, C. F., Lara, Y. J., Cornet, L., François, C., Baurain, D., electromagnetic field on the batch cultivation and nutri- Wilmotte, A., & Javaux, E. J. (2019). Cyanobacteria evolution: tional composition of Spirulina platensis in an air-lift insight from the fossil record. Free radical biology & medicine, photobioreactor. Bioresource Technology, 98, 700–705. 140, 206–223. doi:10.1016/j.freeradbiomed.2019.05.007 doi:10.1016/j.biortech.2006.01.024 Fernie, A. R., & Bauwe, H. (2020). Wasteful, essential, evolu- Lima, G. M., Teixeira, P. C. N., Teixeira, C. M. L. L., tionary stepping stone? The multiple personalities of the Filócomo, D., & Lage, C. L. S. (2018). Influence of spectral photorespiratory pathway. The Plant Journal, 102, 666–677. light quality on the pigment concentrations and biomass doi:https://doi.org/10.1111/tpj.14669 productivity of Arthrospira platensis. Algal Research, 31, Frontasyeva, M. V., Pavlov, S. S., Mosulishvili, L., Kirkesali, E., 157–166. doi:https://doi.org/10.1016/j.algal.2018.02.012 Ginturi, E., & Kuchava, N. (2009). Accumulation of trace Liu, J., Pemberton, B., Lewis, J., Scales, P., & Martin, G. J. O. elements by biological matrix of Spirulina platensis. The (2020). Wastewater treatment using filamentous algae – Journal of the Society of Ecological Chemistry and a review. Bioresource Technology, 298, 122556. doi:https:// Engineering, 16, 277–285. doi.org/10.1016/j.biortech.2019.122556 Frumento, D., Casazza, A. A., Al Arni, S., & Converti, A. Nicolás-Martín, C., Santos-Martín, D., Chinchilla-Sánchez, (2013). Cultivation of Chlorella vulgaris in Tubular M., & Lemon, S. (2020). A global annual optimum tilt Photobioreactors: A Lipid Source for Biodiesel angle model for photovoltaic generation to use in the Production. Biochemical engineering journal, 81, 120–25. absence of local meteorological data. Renewable Energy, doi:10.1016/j.bej.2013.10.011 161, 722–735. doi:10.1016/j.renene.2020.07.098 Ganesh, A. B., Manoharan, P. T., & Suraishkumar, G. K. Nishiyama, Y., Allakhverdiev, S. I., Yamamoto, H., (2007). Responses of the photosynthetic machinery of Hayashi, H., & Murata, N. (2004). Singlet oxygen inhibits Spirulina maxima to induced reactive oxygen species. the repair of photosystem ii by suppressing the translation Biotechnology and Bioengineering, 96, 1191–1198. doi: elongation of the D1 Protein in Synechocystis sp. PCC 6803. https://doi.org/10.1002/bit.21217 Biochemistry-US, 43, 11321–11330. doi:10.1021/bi036178q González-Camejo, J., Viruela, A., Ruano, M. V., Barat, R., Nomsawai, P., de Marsac, N. T., Thomas, J. C., Tanticharoen, M., Seco, A., & Ferrer, J. (2019). Dataset to assess the shadow & Cheevadhanarak, S. (1999). Light regulation of phycobili- effect of an outdoor microalgae culture. Data in Brief, 25, some structure and gene expression in Spirulina platensis C1 104143. doi:10.1016/j.dib.2019.104143 (Arthrospira sp. PCC 9438). Plant & Cell Physiology, 40, 1194– Hoseini, S. M., Almodares, A., Afsharzadeh, S., 1202. doi:10.1093/oxfordjournals.pcp.a029507 Shahriari, A. R., & Montazeri, F. (2014). Growth response Olaizola, M., & Duerr, E. O. (1990). Effects of light intensity of Spirulina platensis PCC9108 to elevated CO levels and and quality on the growth rate and photosynthetic pigment flue gas. Biological Journal of Microorganism, 2, 29–36. content of Spirulina platensis. Journal of Applied Phycology, University of Isfahan, Iran https://www.sid.ir/en/journal/ 2, 97–104. doi:10.1007/BF00023370 ViewPaper.aspx?id=491905 Oliveira, M. A. C. L. D., Monteiro, M., Robbs, P. G., & Ismaiel, M. M. S., El-Ayouty, Y. M., & Piercey-Normore, M. Leite, S. G. F. (1999). Growth and chemical composition (2016). Role of pH on antioxidants production by Spirulina of Spirulina maxima and Spirulina platensis biomass at (Arthrospira) platensis. Brazilian Journal of Microbiology, different temperatures. Aquaculture International, 7, 261– 47, 298–304. doi:10.1016/j.bjm.2016.01.003 275. doi:https://doi.org/10.1023/A:1009233230706 Joshi, S. (2018). Applications of Algae in Cosmetics: An Poonam, S., & Sharma, N. (2017). Industrial and biotechno- Overview. International Journal of Innovative Research in logical applications of algae: A review. Journal of Advances Science, Engineering and Technology, 7, 1269–1278. in Plant Biology, 1, 01–25. doi:https://doi.org/10.14302/ Kazbar, A., Cogne, G., Urbain, B., Marec, H., Le-Gouic, B., issn.2638-4469.japb-17-1534 Tallec, J. . . . Pruvost, J. (2019). Effect of dissolved oxygen Prates, D. D., Radmann, E. M., Duarte, J. H., de Morais, M. G., concentration on microalgal culture in photobioreactors. & Costa, J. A. V. (2018). Spirulina cultivated under different Algal Research, 39, 101432. doi:10.1016/j.algal.2019.101432 light emitting diodes: Enhanced cell growth and phycocya- Kendirlioglu, G., & Cetin, A. K. (2017). Effect of different nin production. Bioresource Technology, 256, 38–43. doi:10. wavelengths of light on growth, pigment content, and 1016/j.biortech.2018.01.122 14 J. L. GAL ET AL. Priyadarshani, I., & Biswajit, R. (2012). Commercial and Srinivasan, T., & Illanjiam, S. (2021b). Optimization studies of industrial applications of micro algae–a review. Journal of multistress spirulina isolated from marine water. Applied Algal Biomass Utilization, 3, 89–100. Ecological and Environmental Sciences, 9, 76–78. Rasala, B. A., & Mayfield, S. P. (2015). Photosynthetic biomanu- Stanley, J. G., & Jones, J. B. (1976). Feeding Algae to Fish. facturing in green algae; production of recombinant proteins Aquaculture, 7, 219–23. doi:10.1016/0044-8486(76)90140-X for industrial, nutritional, and medical uses. Photosynthesis Tacon, A. G. J., & Metian, M. (2008). Global overview on the Research, 123, 227–239. doi:10.1007/s11120-014-9994-7 use of fish meal and fish oil in industrially compounded Rizzo, R.F., Santos, B.D., Castro, G.F., Passos, T.S., aquafeeds: Trends and future prospects. Aquaculture, 285, Nascimento, M.D., Guerra, H.D. . . . Lima-Araújo, K.G. 146–158. doi:10.1016/j.aquaculture.2008.08.015 (2015). Production of Phycobiliproteins by Arthrospira Tayebati, H., Shariati, F. P., Soltani, N., & Tehrani, H. S. (2020). platensis under different light conditions for application The effect of different light variables on spirulina growth and in food products. Food Science and Technology -Brazil, 35, its component. Iranian International Conference. Chemical 247–252. doi:10.1590/1678-457x.6463 Engineering Congress, Exhibition 11. https://www.research Sforza, E., Pastore, M., Franke, S. M., & Barbera, E. (2020). gate.net/profile/HaniehTayebati/publication/345775863 Modeling the Oxygen Inhibition in Microalgae: An Thaweedet, C., Traichaiyaporn, S., & Deming, R. L. (2012). Experimental Approach Based on Photorespirometry. New Effect of light quality on biomass and pigment production biotechnology, 59, 26–32. doi:10.1016/j.nbt.2020.06.003 in photoautotrophic and mixotrophic cultures of Shahi, K. A., Behnaz, E. K., Dehghanian, Z., Pandey, B., Spirulina platensis. Journal of Agricultural Technology, 8, Lajayer, A., Price, G. W., & Astatkie, T. (2022). Removal 1593–1604. of organic and inorganic contaminants from the air, soil, Torzillo, G., & Vonshak, A. (2013). Environmental Stress and water by algae. Environmental Science and Pollution Physiology with Reference to Mass Cultures. Handbook of Research. 06/2022 online. doi: 10.1007/s11356-022-21283-x microalgal culture: applied phycology and biotechnology (pp. Sheehan, J., Dunahay, T. G., Benemann, J. R., Roessler, P. G., 901–13). New Jersey, USA: Blackwell Publishing Ltd. & Weissman, J. C. (1998). A Look back at the U. doi:10.1002/9781118567166.ch6 S. department of energy’s aquatic species program—biodie- Uslu, L. H., Oya, I., Sayin, S., Durmaz, Y., Göksan, T., & sel from algae. National Renewable Energy Laboratory, Gökpinar, Ş. (2009). The effect of temperature on protein Golden, CO, US Department of Energy, Close-out Report. and amino acid composition of Spirulina platensis. Journal https://www.nrel.gov/docs/legosti/fy98/24190.pdf of Fisheries and Aquatic Sciences, 26, 139–142. Shigesada, N., & Okubo, A. (1981). Analysis of the self-shad- Vonshak, A., Torzillo, G., Accolla, P., & Tomaselli, L. (1996). ing effect on algal vertical distribution in natural waters. Light and oxygen stress in Spirulina platensis (cyanobac- Journal of Mathematical Biology, 12, 311–326. doi:10.1007/ teria) grown outdoors in tubular reactors. Physiologia BF00276919 Plantarum, 97, 175–179. doi:10.1111/j.1399-3054.1996. Soni, R. A., Sudhakar, K., & Rana, R. S. (2017). Spirulina – from tb00494.x growth to nutritional product: A review. Trends Food Science Wang, C. Y., Fu, C.-C., & Liu, Y.-C. (2007). Effects of using & Technology, 69, 157–171. doi:10.1016/j.tifs.2017.09.010 light-emitting diodes on the cultivation of Spirulina platen- Soni, R. A., Sudhakar, K., & Rana, R. S. (2019). Comparative sis. Biochemical Engineering Journal, 37, 21–25. doi:10. study on the growth performance of Spirulina platensis on 1016/j.bej.2007.03.004 modifying culture medium. Energy Reports, 5, 327–336. Warman, P. R., & Mayhew, W. J. (1979). Effect of reflective doi:10.1016/j.egyr.2019.02.009 surfaces on a greenhouse lettuce crop. Canada. https:// Srinivasan, T., & Illanjiam, S. (2021a). Extraction and purifi - www.osti.gov/etdeweb/biblio/8487983 cation of phycocyanin and their radical- scavenging activity Webber, A. N., & Lubitz, W. (2001). P700: The primary from multi - stress spirulina isolated from marine water. electron donor of photosystem I. Biochimica Et Biophysica Applied Ecological and Environmental Sciences, 9, 73–75. Acta, 1507, 61–79. doi:10.1016/S0005-2728(01)00198-0
Journal
Applied Phycology
– Taylor & Francis
Published: Dec 31, 2023
Keywords: AlgaTube™; Arthrospira; biomass; cyanobacteria; feed; fertilizer; nutrition; photobioreactor; wastewater; Spirulina
