Extraction of phenolic fraction from guava seeds (Psidium guajava L.) using supercritical carbon dioxide and co-solvents

In this work the supercritical fluid extraction (SFE) with carbon dioxide (CO₂) and with ethyl acetate (EtAc) and ethanol (EtOH) as co-solvents was applied to obtain the phenolic fraction from guava seeds (Psidium guajava L.). The extraction was explored at various operating conditions, using 10, 20 and 30 MPa and 40, 50 and 60 °C. The use of EtAc and EtOH as co-solvents in SFE was also studied. The supercritical process was compared with traditional techniques such as Soxhlet extraction using EtAc and EtOH as solvents. The quality of the different extracts, obtained using SFE and Soxhlet methods and different solvents, was evaluated through the antioxidant activity, obtained by the collection methods of scavenging DPPH and bleaching of β-carotene, and also through the total phenolic content (TPC) of the samples, by the Folin-Ciocalteu method. The antioxidant potential indicates the use of ethanol as co-solvent as the best modifier in SFE, used in concentration of 10% (w/w) at 50 °C and 30 MPa. The quality of the extracts obtained by SFE with EtOH varied with the operating conditions of temperature and pressure, with higher values obtained at 10 and 20 MPa for TPC results and also antioxidant methods. The process yield of the phenolic fraction was also evaluated for all the extraction procedures studied (SFE and Soxhlet), with results varying from 0.380 to 1.738% (w/w).     

Comparison of different extraction methods for the extraction of major bioactive flavonoid compounds from spearmint (Mentha spicata L.) leaves

Different bioactive flavonoid compounds including catechin, epicatechin, rutin, myricetin, luteolin, apigenin and naringenin were obtained from spearmint (Mentha spicata L.) leaves by using conventional soxhlet extraction (CSE) and supercritical carbon dioxide (SC-CO₂) extraction at different extraction schemes and parameters. The effect of different parameters such as temperature (40, 50 and 60 °C), pressure (100, 200 and 300 bar) and dynamic extraction time (30, 60 and 90 min) on the supercritical carbon dioxide (SC-CO₂) extraction of spearmint flavonoids was investigated using full factorial arrangement in a completely randomized design (CRD). The extracts of spearmint leaves obtained by CSE and optimal SC-CO₂ extraction conditions were further analyzed by high performance liquid chromatography (HPLC) to identify and quantify major bioactive flavonoid compounds profile. Comparable results were obtained by optimum SC-CO₂ extraction condition (60 °C, 200 bar, 60 min) and 70% ethanol soxhlet extraction. As revealed by the results, soxhlet extraction had a higher crude extract yield (257.67 mg/g) comparing to the SC-CO₂ extraction (60.57 mg/g). Supercritical carbon dioxide extract (optimum condition) was found to have more main flavonoid compounds (seven bioactive flavonoids) with high concentration comparing to the 70% ethanol soxhlet extraction (five bioactive flavonoids). Therefore, SC-CO₂ extraction is considered as an alternative process compared to the CSE for obtaining the bioactive flavonoid compounds with high concentration from spearmint leaves.     

Brazilian Ginseng extraction via LPSE and SFE: Global yields, extraction kinetics, chemical composition and antioxidant activity

Brazilian Ginseng extracts of two species, Pfaffia paniculata and Pfaffia glomerata, were obtained by supercritical fluid extraction (SFE) with CO₂ and by low-pressure solvent extraction (LPSE) with methanol, hexane and ethanol. The SFE assays were conducted at pressures of 100, 200 and 300 bar, and temperatures of 30 and 50 °C. The qualitative chemical compositions of the extracts were determined by thin layer chromatography (TLC). One of the active principles of interest from P. glomerata extract, β-ecdysone, was identified and quantified by HPLC. The antioxidant activities of Brazilian Ginseng extracts were determined by the coupled reaction of linolenic acid and β-carotene. For P. paniculata, the highest SFE yield was obtained at 200 bar/50 °C (0.22%, dry basis—d.b.), while the best extraction condition for P. glomerata was obtained at 200 bar/30 °C (0.18%, d.b.). The higher extract yields obtained by LPSE were 2.0% and 5.8% (w/w, d.b.) for P. paniculata and P. glomerata, respectively, both obtained with methanol as extraction solvent. From the overall extraction curve of P. glomerata, it was possible to obtain the kinetic parameters of extraction; the duration of the CER (constant extraction rate) period was determined as 134 min. The TLC plates showed the possible presence of flavonoids in the ethanolic extract for both Pfaffia species. The antioxidant activity analysis detected that LPSE extracts had higher activity than SFE extracts.     

Supercritical fluid extraction of flavonoids from Maydis stigma and its nitrite-scavenging ability

Supercritical fluid carbon dioxide (SF-CO₂) extraction (SFE) of flavonoids from Maydis stigma and its nitrite-scavenging ability were investigated. The effects of extraction time, particle size and co-solvent composition in terms of water content in ethanol were first optimized. Then, a Box-Behnken design combined with response surface methodology (RSM) was employed to study the effects of three independent variables (temperature, pressure and co-solvent amount) on the extraction yield of flavonoids. A maximal extraction yield of flavonoids of approximately 4.24 mg/g of M. stigma by SFE was obtained under optimal conditions (a temperature of 50.88 °C, a pressure of 41.80 MPa, a co-solvent amount of 2.488 mL/g and an extraction time of 120 min with 0.4-mm particle sizes and 20% aqueous ethanol as the co-solvent). Furthermore, the nitrite-scavenging ability of the flavonoid-enriched SFE extracts was assessed using the Griess reagent. The flavonoid-enriched SFE extracts exhibited the highest scavenging ability on nitrite (88.1 ± 3.04%) at the concentration of 500 μg/mL and at pH 3.0. The nitrite-scavenging ability of the extracts appeared to be concentration dependent but negatively correlated with the pH.     

Radical-scavenging activity of extracts from Cordia verbenacea DC obtained by different methods

The possibility of using the leaves of Cordia verbenacea as a new source of natural antioxidant compounds was investigated. In the present work, extracts from C. verbenacea were obtained using different extraction methods: supercritical fluid extraction (SFE), Soxhlet (SE), hydrodistillation and maceration, with the objective to evaluate the methods in terms of yield and antioxidant potential. The high-pressure technique was applied using pure CO₂ and CO₂ with co-solvent at different temperatures and pressures (30, 40 and 50 °C and 100, 200, and 300 bar). Organic solvents with different polarities were used to obtain extracts by low-pressure extraction processes. The extracts were evaluated according to their antioxidant activity using total phenolic content, scavenging abilities on DPPH radical, total antioxidant activities (ABTS^•+), superoxide anion radical-scavenging (O₂^•) and protection against lipid peroxidation in vitro (LPO). Ethyl acetate fraction obtained by maceration and extract isolated by SE using 25% aqueous mixture of ethanol possessed the highest scavenger activity against DPPH radical (IC₅₀ = 9.2 ± 0.4 μg/ml, IC₅₀ = 27.4 ± 0.1 μg/ml, respectively). The SFE with 8% ethanol as a co-solvent produced extracts with distinguished increase in the antioxidant activity. The Soxhlet extract with ethyl acetate exhibited a strong reduction of lipid peroxidation (IC₅₀ = 209 ± 3 μg/ml) value comparable to the standard rutin (IC₅₀ = 203 ± 2 μg/ml). The results indicate that extracts of C. verbenacea have important potential as a source of bioactive compounds with antioxidant activity.     

Fractionation of thyme (Thymus vulgaris L.) by supercritical fluid extraction and chromatography

Supercritical fluid chromatography (SFC) was employed to fractionate thyme (Thymus vulgaris L.) extracts, which were obtained by supercritical carbon dioxide extraction of thyme leaves. First, different supercritical extracts were produced at 313 K and at different pressures (15, 30 and 40 MPa). Thymol, a monocyclic terpenoid with recognized antiseptic, analgesic and anti-inflammatory properties, was identified and quantified in the different samples by GC-MS. Then, the supercritical extracts were fractionated by semi-preparative SFC, and different conditions such as pressure, temperature and amount of cosolvent (ethanol) employed were studied. Around a two fold increase of thymol was achieved at 15 MPa, 50 °C and 3% ethanol cosolvent, recovering 97% of the monocyclic terpenoid extracted.     

A green separation process for recovery of healthy oil from pumpkin seed

In this study, pumpkin seeds, called as “Ürgüp Sivrisi” and grown in Cappadocia region, were used as plant materials because of high aroma contents. In the supercritical fluid extraction of pumpkin seed oil, the effect of main process parameters as the particle size (250-2360 μm), the volumetric flow rate of supercritical solvent (0.06-0.30 L/h), the operating pressure (20-50 MPa), the operating temperature (40-70 °C), the type of entrainer (ethanol and n-hexane) and those concentrations (0-10 vol.%) on the extraction yield, the oil solubility and the initial extraction rate were investigated. A cross-over effect for the extraction of pumpkin seed oil using supercritical CO₂ was determined at the operating pressure of 20-30 MPa. The maximum extraction yield obtained with entrainer free was reached 0.50 g oil/g dry seed at 600-1180 μm, 0.12 L/h, 50 MPa and 70 °C for the operation time of 5 h. The maximum extraction yield obtained with ethanol as an entrainer in the experiments was reached 0.54 g oil/g dry seed at the conditions of 600-1180 μm, 0.12 L/h, 30 MPa, 40 °C and 8 vol.% for the operating time of 2 h. The oil compositions were determined by gas chromatography analysis and the results showed that the compositions of pumpkin seed oil which were obtained by means of organic solvent extraction and supercritical fluid extraction were similar. The average oil compositions determined as 9.3 (±0.43)% palmitic acid, 7.5 (±0.6)% stearic acid, 32.3 (±0.6)% oleic acid, 48.1 (±0.6)% linoleic acid and 0.7 (±0.3)% linolenic acid. The morphological changes in the seeds were determined by the scanning electron microscopy analysis.     

Extraction and identification of proanthocyanidins from grape seed (Vitis Vinifera) using supercritical carbon dioxide

In this study, supercritical carbon dioxide extraction of proantocyanidins (PRCs) was performed and the effect of different pressure, temperature and ethanol percentage was investigated. High performance liquid chromatography was used for the analysis of the compounds and it was found that the most effective parameter on the extraction was the amount of the ethanol percentage. Each compound was extracted from grape seeds at their maximum level when different parameters were used which was probably because of their different polarities. Gallic acid (GA), epigallocatechin (EGC) and epigallocatechingallate (EGCG) were extracted at their maximum level when the 300 bar 50 °C and 20% of ethanol was used. The maximum amount of catechin (CT) and epicatechin (ECT) were obtained when 300 bar 30 °C and 20% of ethanol was used for extraction, and 250 bar, 30 °C and 15% of ethanol was needed to extract highest amount of epicatechingallate (ECG).     

Kinetics of ultrasound assisted extraction of anthocyanins from Aronia melanocarpa (black chokeberry) wastes

Black chokeberry fruits are very rich in antioxidant phenolics. After juice extraction, the wastes of pressed berries (juice production by-products) still contain important amount of phenolic species and namely anthocyanins. Ultrasound assisted extraction (UAE) from such wastes was studied. The influence of extraction time (0–240 min), temperature (20–70 °C), solvent composition (0–50% ethanol in water) and ultrasound power (0–100 W) on anthocyanins and polyphenols yields and on antioxidant capacity of the extracts was studied. High temperature and ethanol content in the solvent improved greatly the extraction yields. Ultrasounds improved mainly the extraction kinetics. Ultrasound effect was higher in the beginning of extraction process and at low temperatures. Suitability of UAE for preparation of antioxidant-rich plant extracts at reduced time and energy was confirmed obtaining high extraction efficiencies and high antioxidant capacities of Aronia extracts. To optimize extraction conditions a kinetic mathematical model, based on Peleg's equation and considering also the rate of anthocyanins thermal degradation, was proposed. The influences of extraction time, temperature, solvent composition and ultrasound power on polyphenols and anthocyanins yields and on antioxidant capacity were considered. The optimal conditions for anthocyanins extraction predicted by this model were experimentally validated.     

Electrochemical behaviour of ZrO2 sol-gel pre-treatments on AA6060 aluminium alloy

ZrO₂ pre-treatments applied with the sol-gel technique are a possible replacement of chromium based pre-treatments on aluminium alloys. The thickness and homogeneity of the films deposited on AA6060 alloy are strongly related to the process parameters like preparation of the surface, number of dips and thermal treatment of the film.ZrO₂ films were prepared using the dip-coating technique in sol obtained from metal-organic precursors in an organic solvent (0.1 M Zr(OBuⁿ)₄ in anhydrous n-butanol with addition of acetic acid as complexing agent). Different layers were applied on AA6060 changing number of dips and thermal treatment (150 °C for 1 h or 250 °C for 4 min). The typical thickness of the deposited layers was in the range 70-180 nm depending on process parameters. The electrochemical behaviour of the pre-treated alloy in diluted Harrison solution (0.05 wt% NaCl + 0.35 wt% (NH₄)₂SO₄) was investigated by means of potentiodynamic polarization, open circuit potential measurements, and electrochemical impedance spectroscopy. In addition, the electrochemical behaviour of ZrO₂ sol-gel films was compared with that of chromatized AA6060 and fluotitanated/fluozirconated AA6060. In order to evaluate the adhesion properties of the films, ZrO₂ pre-treated AA6060, chromatized AA6060 and fluotitanated/fluozirconated AA6060 were painted with a polyester resin and subjected to thermal cycles in 0.05 wt% NaCl. Each thermal cycle consisted of heating the samples at 90 °C, permanence at 90 °C for 6 h, cooling at room temperature and permanence at room temperature for 18 h. Impedance measurements were performed at the end of each cycle.Potentiodynamic polarization curves and impedance spectra indicate that ZrO₂ pre-treatments have similar barrier properties to those of chromatized AA6060. However, no self-healing ability is observed for ZrO₂ films.The barrier properties of ZrO₂ films are strongly dependent on process parameters. In particular, the number of dips determines the amount of defects in the film and its homogeneity. The electrochemical behaviour strongly improves increasing the number of dips in the deposition bath. Thermal aging cycles evidence good adhesion properties for ZrO₂ pre-treatments.     

Supercritical fluid extraction from dried banana peel (Musa spp., genomic group AAB): Extraction yield, mathematical modeling, economical analysis and phase equilibria

Supercritical fluid extraction from dried banana peel (Musa spp., subgroup Prata, genomic group AAB, popularly known in Brazil as Enxerto) was studied. The aspects investigated were: overall extraction curve (OEC), mass transfer modeling of the yield curves, economical analysis of the process and phase equilibrium data for the pseudo-ternary system of banana peel extract, carbon dioxide and ethanol. The extraction operating conditions evaluated were: pressure ranging from 100 bar to 300 bar, temperature from 40 to 50 °C and constant solvent flow rate of 5.0 gCO₂/min. Experimental extraction data were correlated using three kinetic models based on mass transfer equations (logistic, diffusion and Esquível models). Phase equilibrium measurements were performed using pressure from 64.9 bar to 239.9 bar and mass fraction of supercritical extract from 0.52 to 3.55 wt%. Yield results ranged from 0.6 to 6.9% d.b. (dry basis). The lowest deviation between experimental and correlated data was obtained by the Logistic model, except for the curve at 300 bar and 40 °C which was best represented by the Esquível model. The economical analysis identified the possibility to apply the supercritical fluids to obtain extracts from banana peel in an industrial scale. Phase equilibrium for the supercritical extract from banana peel with carbon dioxide modified by ethanol exhibited liquid-liquid, vapor-liquid (bubble point) and vapor-liquid-liquid phase transitions. A crossover phenomenon for the systems evaluated was observed for pressures between 200 bar and 240 bar, for both groups of assays, i.e., supercritical extraction and phase equilibrium.     

Antioxidant Activity and Total Phenolics of Propolis from the Basque Country (Northeastern Spain)

The purpose of this study was to evaluate the antioxidant activity (AA) of 19 propolis extracts prepared in different solvent (ethanol and propylene glycol). It was observed that all the samples tested had AA, although results varied considerably between extracts, i.e. 420–1,430 μmol Trolox/g (ABTS), 108–291 mg ascorbic acid/g (DPPH), and 1,573–4,669 μmol iron⁺⁺ sulfate/g (FRAP). The ethanol may enhance the potency of the AA, and the correlation coefficient between total phenolic content (TPC) (200–340 mg/g propolis extracts) and AA was statistically significant. Total flavonoids ranged from 72 to 161 mg/g propolis extracts. The results indicate that TPC and flavonoids contributed to AA.     

A food grade approach for the isolation of major alkylresorcinols (ARs) from rye bran applying tailored supercritical carbon dioxide (scCO2) extraction combined with HPLC

Tailored supercritical carbon dioxide (scCO₂) extraction of alkylresorcinols (ARs) from rye bran resulting in pre-purification of ARs already during extraction process. The pre-purification was achieved by splitting the extraction process in 2 steps._. In both steps, the CO₂ pressure was set at 25 MPa with a flow rate of 10 g/min. The first step was carried out at 70 °C, 0.06% of ethanolic co-solvent for 2 h, followed by the second step at 45 °C using 10% of ethanolic co-solvent for 4 h, resulting in a pre-purified ARs extract. From the pre-purified scCO₂ extract, chromatographed on C8 column, were obtained pure ARs homologues of C17, C19 and C21 (68% of total) eluting separately in a linear gradient of ethanol. Additionally, a rapid method for the isolation of high purified pools of ARs homologues using a disposable solid phase extraction SPE-C18 column and a step gradient of ethanol was developed.     

Optimization of supercritical carbon dioxide extraction of Passiflora seed oil

This study investigates extraction of Passiflora seed oil by using supercritical carbon dioxide. Artificial neural network (ANN) and response surface methodology (RSM) were applied for modeling and the prediction of the oil extraction yield. Moreover, process optimization were carried out by using both methods to predict the best operating conditions, which resulted in the maximum extraction yield of the Passiflora seed oil. The maximum extraction yield of Passiflora seed oil was estimated by ANN to be 26.55% under the operational conditions of temperature 56.5 °C, pressure 23.3 MPa, and the extraction time 3.72 h; whereas the optimum oil extraction yield was 25.76% applying the operational circumstances of temperature 55.9 °C, pressure 25.8 MPa, and the extraction time 3.95 h by RSM method. In addition, mean-squared-error (MSE) and relative error methods were utilized to compare the predicted values of the oil extraction yield obtained from both models with the experimental data. The results of the comparison reveal the superiority of ANN model compared to RSM model.     

Supercritical CO2 extraction of Helichrysum italicum: Influence of CO2 density and moisture content of plant material

Kinetics and selectivity of supercritical carbon dioxide (SC CO₂) extraction of Helichrysum italicum flowers were analyzed at pressures in the range of 10-20 MPa and temperatures of 40 °C and 60 °C (density of SC CO₂ from 290 to 841 kg/m³) and also at 10 MPa and 40 °C using flowers with different moisture contents (10.5% and 28.4%). Increased moisture content of H. italicum flowers resulted in enchased solubility of solute enabling decrease of SC CO₂ consumption necessary for achieving desired extraction yield. The most abundant compounds in the supercritical extracts are sesquiterpenes and waxes while monoterpenes and sesquiterpenes are the main constituents of essential oil obtained by hydrodistillation. The optimal set of working parameters with respect to extraction yield, SC CO₂ consumption and chemical composition of extract were defined related to moisture content of raw material and SC CO₂ density.     

Optimization of process variables for essential oil components from Satureja hortensis by supercritical fluid extraction using Box-Behnken experimental design

The oil and extracts of Satureja hortensis cultivated in Iran were extracted using supercritical carbon dioxide and hydrodistillation method. The oil and extracts were analyzed by GC-FID and GC/MS. The compounds were identified according to their retention indices and mass spectra (EI, 70 eV). The effects of various parameters such as pressure, temperature, percent of modifier (methanol) and extraction time, were investigated by a fractional factorial design (2^4-1) to determine the significant parameters and their interactions. The results showed that the pressure, temperature and percent of modifier are significant (p < 0.05), but the extraction time was found to be insignificant. The response surface methodology (RSM), based on Box-Behnken design was employed to obtain the optimum conditions of the significant parameters (pressure, temperature and percent of modifier). The optimal conditions could be obtained at a pressure of 35.0 MPa, temperature of 72.6 °C, and 8.6% (v/v) for methanol. The main extracted components using SFE were γ-Terpinene (35.5%), Thymol (18.2%) and Carvacrol (29.7%).     

Negative pressure cavitation accelerated processing for extraction of main bioactive flavonoids from Radix Scutellariae

To enhance the extraction efficiency and reduce the energy consumption, an emerging technology named negative pressure cavitation extraction (NPCE) has been shown to be a feasible option for the extraction of bioactive compounds in agricultural crops and medicinal plants. Meanwhile, it can be applied at the pilot scale as a manufacturing process for edible and medicinal plants. Currently, NPCE was proposed for extraction of baicalin, wogonoside, baicalein and wogonin from Radix Scutellariae on the basis of a central composite design (CCD) and response surface methodology (RSM). With proper optimization (80 mesh of particle size, 40 mL/g of liquid/solid ratio, 75% aqueous ethanol as extraction solvent, 60 min extraction time and −0.07 MPa vacuum degree), NPCE was observed to have good extraction efficiency compared with other conventional extraction methods. Furthermore, the antioxidant activities of crude extracts with different extraction methods were assessed by the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical-scavenging assay. Our results showed that NPCE extract possessed better antioxidant activity with IC₅₀ value of 3.24 μg/mL compared with the UAE, HRE and SE extracts with IC₅₀ values of 7.85, 12.14 and 11.44 μg/mL, respectively.     

Countercurrent supercritical fluid extraction of grape-spirit

A procedure for the recovery of aromatic extracts from raw grape-spirit (alcohol strength 73 vol.%) by means of a countercurrent supercritical fluid extraction (CC-SFE) on a pilot plant scale is presented. The effect of the solvent-to-feed ratio at 7, 8, 10, and 20 kg_CO2/kg_Feed on the extraction of flavor compounds, operating at constant pressure of 100 bar and extraction temperature of 50 °C have been studied. Increasing the solvent-to-feed ratio from 7 to 20 kg_CO2/kg_Feed, the total volatile compounds yield decreases from 64 to 5% and that of ethanol from 85 to 49%. The highest concentration of volatile compounds has been obtained at 7 and 8 kg_CO2/kg_Feed solvent-to-feed ratio. Mixtures containing grape-spirit (alcohol strength 40 vol.%) added with 30% (v/v) of different extracts have been tested and judged by a panel. Sensory evaluation indicated as preferred the mixtures added with the extracts obtained at lowest solvent-to-feed ratio.     

Effect of annealing on the properties of nanostructured CuO thin films for enhanced ethanol sensitivity

Copper oxide (CuO) thin films were grown on glass substrates by low cost spray pyrolysis technique for three different molar concentrations (0.05 M, 0.10 M and 0.15 M), at a substrate temperature of 350 °C, and subsequently annealed at 400 °C for 2 h. The effects of precursor concentration and annealing on the structural, electrical and morphological properties of the crystallized films were investigated. X-ray diffractograms of the films showed the formation of single phase CuO with tenorite structure. The electrical properties of the films like carrier concentration, Hall co-efficient (R_H), mobility and conductivity were studied from Hall effect measurements. The positive values of R_H confirmed the p-type conductivity of the films. Resistivity decreased drastically by two orders of magnitude for the annealed films. The microstructures characterized by a scanning electron microscope for 0.15 M concentration of the precursor revealed that the morphology of the films was substantially affected by annealing. The film surface revealed uniformly distributed cluster of peanut shaped grains after annealing. The response of the as deposited and annealed CuO sensor to low concentration of ethanol (10 ppm) was compared. The annealed CuO film showed higher sensor response than the as-deposited CuO film did. The result suggested that annealing causes significant effect on the sensing performance of CuO to ethanol.     

Microencapsulation of Morinda citrifolia L. extract by spray-drying

Microcapsules containing Morinda citrifolia L. microparticles were produced by a spray-drying technique using various proportions of κ-carrageenan and maltodextrin as the binding materials. In this work, the effects of spray-drying on the encapsulation yield, particle size, moisture content, DPPH scavenging activity, total phenolic content and total flavonoid content of the bioactive components of M. citrifolia L. were determined for different volume ratios in the inlet air temperature range of 90–140 °C. The results showed that the percentage of 2,2-diphenyl picrylhydrazyl (DPPH) scavenging activity of the spray-dried powder was the highest for the 1:2 ratio (volume ratio of M. citrifolia L. extract to additive solution) at 90 °C, with maltodextrin at a concentration of 33 mg/ml. The results also showed that the microcapsules had a regular spherical shape. The spray-dried M. citrifolia fruit extract showed high antioxidant activity (28.36% DPPH activity), thus suggesting that it might be useful as a food additive and/or ingredient under the above optimum operating conditions.