Degradation Of PHB Research Articles

One-Pot Biosynthesis of Acetone from Waste Poly(hydroxybutyrate).

The plastic waste crisis is catalyzing change across the plastics life cycle. Central to this is increased production and application of bioplastics and biodegradable plastics. In particular, poly(hydroxybutyrate) (PHB) is a biodegradable bioplastic that can be produced from various renewable and waste feedstocks and is a promising alternative to some petrochemical-derived and non-biodegradable plastics. Despite its advantages, PHB biodegradation depends on environmental conditions, and the effects of degradation into microplastics, oligomers, and the 3-hydroxybutyrate (3-HB) monomer on soil microbiomes are unknown. We hypothesized that the ease of PHB biodegradation renders this next-generation plastic an ideal feedstock for microbial recycling into platform chemicals currently produced from fossil fuels. To demonstrate this, we report the one-pot degradation and recycling of PHB into acetone using a single strain of engineered Escherichia coli. Following strain development and initial bioprocess optimization, we report maximum titers of 123 mM acetone (7 g/L) from commercial PHB granules after 24 h fermentation at 30 °C. We further report biorecycling of an authentic sample of post-consumer PHB waste at a preparative scale. This is the first demonstration of biological recycling of PHB into a second-generation chemical, and it demonstrates next-generation plastic waste as a novel feedstock for the circular bioeconomy.

3D Filaments Based on Polyhydroxy Butyrate-Micronized Bacterial Cellulose for Tissue Engineering Applications.

In this work, scaffolds based on poly(hydroxybutyrate) (PHB) and micronized bacterial cellulose (BC) were produced through 3D printing. Filaments for the printing were obtained by varying the percentage of micronized BC (0.25, 0.50, 1.00, and 2.00%) inserted in relation to the PHB matrix. Despite the varying concentrations of BC, the biocomposite filaments predominantly contained PHB functional groups, as Fourier transform infrared spectroscopy (FTIR) demonstrated. Thermogravimetric analyses (i.e., TG and DTG) of the filaments showed that the peak temperature (Tpeak) of PHB degradation decreased as the concentration of BC increased, with the lowest being 248 °C, referring to the biocomposite filament PHB/2.0% BC, which has the highest concentration of BC. Although there was a variation in the thermal behavior of the filaments, it was not significant enough to make printing impossible, considering that the PHB melting temperature was 170 °C. Biological assays indicated the non-cytotoxicity of scaffolds and the provision of cell anchorage sites. The results obtained in this research open up new paths for the application of this innovation in tissue engineering.

Poly(3-hydroxybutyrate) Degradation by Bacillus infantis sp. Isolated from Soil and Identification of phaZ and bdhA Expressing PHB Depolymerase.

Poly(3-hydroxybutyrate) (PHB) is a biodegradable and biocompatible bioplastic. Effective PHB degradation in nutrient-poor environments is required for industrial and practical applications of PHB. To screen for PHB-degrading strains, PHB double-layer plates were prepared and three new Bacillus infantis species with PHB-degrading ability were isolated from the soil. In addition, phaZ and bdhA of all isolated B. infantis were confirmed using a Bacillus sp. universal primer set and established polymerase chain reaction conditions. To evaluate the effective PHB degradation ability under nutrient-deficient conditions, PHB film degradation was performed in mineral medium, resulting in a PHB degradation rate of 98.71% for B. infantis PD3, which was confirmed in 5 d. Physical changes in the degraded PHB films were analyzed. The decrease in molecular weight due to biodegradation was confirmed using gel permeation chromatography and surface erosion of the PHB film was observed using scanning electron microscopy. To the best of our knowledge, this is the first study on B. infantis showing its excellent PHB degradation ability and is expected to contribute to PHB commercialization and industrial composting.

Active microbial communities during biodegradation of biodegradable plastics by mesophilic and thermophilic anaerobic digestion.

Biodegradable plastics, if they are not properly managed at their end-of-life, can have the same hazardous environmental consequences as conventional plastics. This study investigates the treatment of the main biodegradable plastics under mesophilic and thermophilic anaerobic digestion using biochemical methane potential test and the microorganisms involved in the process using amplicon sequencing of the 16S rRNA. Here we showed that, only PHB and TPS undergone important and rapid biodegradation under mesophilic condition (38°C). By contrast, PCL and PLA exhibited very low biodegradation rate as 500 days were required to reach the ultimate methane yield. Little or no degradation occurred for PBAT and PBS at 38°C. Under thermophilic conditions (58°C), TPS, PHB, and PLA reached high levels of biodegradation in a relatively short period (< 100 d). While PBS, PBAT, and PCL could not be converted into methane at 58°C. PHB degraders (Enterobacter and Cupriavidus) and lactate-utilizing bacteria (Moorella and Tepidimicrobium) appeared to play an important role in the PHB and PLA degradation, respectively. This work not only provides crucial data on the anaerobic digestion of the main biodegradable plastics but also enriches the understanding of the microorganisms involved in this process, which are of great importance for future development of the treatment of biodegradable plastics in anaerobic digestion systems.

Polyhydroxybutyrate-Natural Fiber Reinforcement Biocomposite Production and Their Biological Recyclability through Anaerobic Digestion

The existing recycling methods of PHA-based material are ineffective in terms of increasing resource efficiency and the production of high value end-of-life products. Therefore, in this study, a novel approach of acidogenic fermentation was proposed to recycle PHB-based composites reinforced with natural fibers such as cellulose, chitin, chitosan, orange waste, sawdust, soy protein, and starch. The inclusion of cellulose, chitosan, and sawdust improved the impact properties of the composites while other fillers had various effects on the mechanical properties. These three composites and neat PHB were subsequently subjected to biological degradation via acidogenic digestion to determine the possibility of converting PHB-based composites into volatile fatty acids (VFAs). Two different pH levels of 6 and 10 were applied to assess the effect of pH on the bioconversion and inhibition of the methanogenesis. The results showed promising PHB degradation, contributing to considerable VFA production of 2.5 g/L at pH 6 after 47 days. At pH 6, the presence of the natural fibers in the biocomposites promoted the degradation rate. On the contrary, pH 10 proved to be more suitable for the degradation of the fibers. The VFA which is produced can be recirculated into PHB production, fitting with the concept of a circulating bioeconomy.

Polyhydroxybutyrate degradation by biocatalyst of municipal sludge water and degradation efficacy in sequencing batch biofilm reactor

The main aim of the study was to degrade poly-β-hydroxybutyrate (P(3HB)) in the sequencing batch biofilm reactor (SBBR) using biocatalyst. Enrichment method was used for the isolation of P(3HB) degrading bacteria. These bacterial strains were isolated from the wastewater sludge sample treated with P(3HB) sheets. A total of 75 bacteria were isolated after 60 days of incubation. The zone of clearance varied between 12 ± 1 mm and 19 ± 2 mm. Two bacterial strains (Nitrobacter vulgaris SW1 and Pseudomonas aeruginosa KS10) showed rapid PHB degradation activity on agar plates. Plate screening experiments confirmed PHB degrading ability of P. aeruginosa KS10 and N. vulgaris SW1. Biodegrading potential improved after 72 h fermentation period. The bacteria produced depolymerase and enzyme activity was maximum after 72 h. The sequencing batch biofilm reactor (SBBR) co-cultured with N. vulgaris SW1 and P. aeruginosa KS10 was operated to remove PHB from the wastewater. Biofilm in the reactor degraded PHB and the production of polyhydroxybutyrate depolymerase influenced on PHB degradation. Polyhydroxybutyrate degradation improved continuously and maximum degradation (95.6%) was achieved after 8 days. The degradation of biopolymers help to reduce environmental pollution associated with the petroleum based polymers.

Effect of calcium stearate as a lubricant and catalyst on the thermal degradation of poly(3-hydroxybutyrate)

Poly(3-hydroxybutyrate) (PHB) is a promising substitute to petroleum-based polymers in packaging and biomedical applications provided that its melt processability and degradability are improved. A new method to control the properties of PHB by using cheap calcium stearate (CS) as a lubricant and decomposition catalyst in melt-mixed PHB-CS compounds was first used. CS is composed of a metallic cation, which promotes PHB degradation, and a hydrophobic anion that improves the compatibility with PHB and processability. An environmentally friendly melt mixing technique was employed to obtain the PHB-CS compounds. Incorporation of 0.5 or 5 wt% CS reduced the melt viscosity and molecular weight of PHB, decreased the melting temperature with up to 5 °C, the crystallization temperature with more than 25 °C, and the degradation temperature with 15 and 40 °C, respectively. In small amounts (0.05 wt%), CS improved the processability and mechanical properties of PHB. In higher amount (0.5 wt%), CS slightly improved the Young's modulus, reduced the tensile strength and enhanced degradation. A better control of thermal and mechanical properties of PHB is, thus, possible by using different CS amount and processing conditions. These results are relevant for PHB application in the context of the global transition to biodegradable packaging.

Screening, optimization and characterization of poly hydroxy butyrate from fresh water microalgal isolates

ABSTRACT Microalgae, especially Chlorella sp., have been used for centuries as food and currently their biotechnological potential is notable for the presence of several compounds relevant to the market, like PHB. Microalgae are appealing because of the increasing demand for biopolymers like PHB. Therefore, the present study is aimed to determine the efficacy of biomass production and yield of PHB producing microalgae isolated and identified by phylogenetic analysis. The parameter was optimized as pH 7, temperature 30°C, under sunlight with 0.2% of sodium bicarbonate for higher biomass and PHB production. The extracted PHB were characterized through SEM, EDAX, XRD and FT-IR. In GC-MS analysis the major peak represents Benzyl butyl phthalate confirming the polymer of PHB. DSC-TGA, was performed and the melting peak of PHB was found as 342.1°C conferring to thermogravimetric analysis, the range of temperature for rapid thermal degradation of PHB was at 418°C to 420°C with the degradation peaking at 418.2°C the total weight loss within this temperature range was 98.5%. Hence, maximum 80% of PHB was produced from Chlorella sp. in addition to sequestering CO2.

Plasticization of poly(3‐hydroxybutyrate) with triethyl citrate: Thermal and mechanical properties, morphology, and kinetics of crystallization

AbstractPoly(3‐hydroxybutyrate) (PHB), is one important biopolymer and a promising alternative to petroleum‐based plastics. In this article, formulations of PHB and triethyl citrate (TEC) as plasticizer were prepared by melt extrusion. The effect of TEC on the mechanical, thermal, and morphological properties of PHB was investigated by tensile tests, impact resistance, dynamic‐mechanical analysis, differential scanning calorimetry, polarized optical microscopy, and small‐ and wide‐angle X‐ray scattering. TEC acted as an efficient plasticizer for PHB, imparting gradual changes in the properties as the mass fraction of TEC increased. A reduction in the elastic modulus, an increase in the intensity of β relaxation indicated a higher capacity of mechanical energy dissipation for the formulations containing higher mass fractions of TEC. TEC reduced its glass transition and melting temperatures, contributing to the increase of the processing window of the temperature and minimizing thermal degradation of PHB. TEC had a strong influence on the kinetics of crystallization, the morphology of the spherulites, and the crystalline structural parameters, such as long period, crystalline lamella, and interlamellar amorphous region thicknesses. Our study clarifies how the morphology of the PHB crystalline phase evolves in the presence of the plasticizer and with the time of crystallization.

Band Spacing in Poly(3-hydroxybutyrate) and Its Blends with Poly(propylene carbonate): Dependence on Thermal Processing.

The band spherulites grown in neat poly(3-hydroxybutyrate) (PHB) and its blends with poly(propylene carbonate) (PPC) were observed by polarized optical microscopy. For the spherulites in neat PHB, it is evident that the band spacing increases first and then decreases with melting time. As the melting time is within 7 min, the band spacing increases continuously, which should be attributed to increasing mobility of polymer chains or decreasing viscosity of the melt. When the melting time is prolonged, evident thermal degradation of PHB occurs and results in a great deal of noncrystalline fractions, which is similar with addition of miscible amorphous polymers in the melt, and the band spacing decreases accordingly. The thermal degradation of PHB cannot, however, be detected by a thermogravimetric analyzer because of less volatile productions. An evident decrease of molecular weight can be measured by gel permeation chromatography, indicating occurrence of serious degradation. The decrease of crystallization and melting temperature revealed by differential scanning calorimetry (DSC) also prove the thermal degradation. For spherulites in PHB/PPC blends, however, the variation of band spacing differs from that in neat PHB. The band spacing increases continuously when melting time is within 15 min. The crystallization and melting behaviors are not influenced greatly by prolonging melting time in PHB/PPC blends. The variations of Mw for PHB/PPC are slighter than those of the neat PHB and PPC upon heating at 190 °C. Combined with the corresponding DSC results, it is conjectured that blending may prohibit the degradation of PHB to some extent. An intermolecular interaction can be detected between PHB and PPC via Fouriertransform infrared spectra and should help to avoid degradation of PHB to a certain degree. The present results may help widen the applications of PHB and shed some light on understanding the formation mechanism of the band for aliphatic polyester polymers.

A novel, thermotolerant, extracellular PHB depolymerase producer Paenibacillus alvei PHB28 for bioremediation of biodegradable plastics

Abstract Background Poly-β-hydroxybutyrate (PHB) is the most important and versatile class of biodegradable polymers used successfully in the medical, agricultural and industrial field. Idea is to find the novel isolate for degradation of biodegradable plastics that can enhance the bioremediation. Materials and methods Thirty-one PHB and PHB depolymerase enzyme producing isolates out of 80 mesophilic bacteria from Lucknow region were further screened for PHB degradation capability by secreting extracellular PHB depolymerase enzyme in minimal salt media supplemented with PHB (0.15%). Various biodegradable plastic films were tested by soil burial method for weight loss determination. Result 37.3% weight loss has been observed in PHB films when buried under the soil for 45 days in the presence of a novel PHB degrader identified as Paenibacillus alvei PHB28 by 16S rRNA sequencing (GenBank accession number KX886342). These Gram-negative, spore-forming, thermotolerant bacteria produce maximum PHB depolymerase (5.03 U/mL) at 45°C, pH 8.0, with 0.15% substrate concentration when incubated for 96 h with starch (0.1%) and yeast extract (0.01%) as an additional nutrient supplements. Conclusion To the best of our knowledge this is the first report of PHB depolymerase production by P. alvei PHB28 which may contribute successfully to combat plastic pollution and to sustain the green environment.

Effect of clay treatment on the thermal degradation of PHB based nanocomposites

Abstract A detailed understanding of the thermal degradation processes taking place during the melt processing of bio-nanocomposites is crucial in order to increase the processing window of these materials. In this work, the influence of the content of neat clay and modified-clay on the thermal degradation of a biodegradable bacterial poly(3-hydroxybutyrate) (PHB) matrix, was studied. The modified clay consisted in a multi-treated organobentonite, which was first acid-activated, then silylated and further modified by cationic exchange treatment. Thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC) tests were carried out to investigate the thermal behavior of the different nanocomposites as a function of temperature, whereas size exclusion chromatography (SEC) runs were performed to analyze the changes on the molecular weight distribution of the PHB. The obtained results reveal that the organic modifiers of the muti-treated clay promote the thermal degradation process leading to a dramatic decrease in the molecular weight of PHB. It was demonstrated that the degradation mechanism of PHB was not modified by the incorporation of neat clay or modified-clay, and that the process can be well described by the Avrami–Erofeev random nucleation model (m = 4), in which the reaction is controlled by initial random nucleation followed by overlapping growth.

Poly(3-Hydroxybutyrate) (PHB) Polymerase PhaC1 and PHB Depolymerase PhaZa1 of Ralstonia eutropha Are Phosphorylated In Vivo.

In this study, we screened poly(3-hydroxybutyrate) (PHB) synthase PhaC1 and PHB depolymerase PhaZa1 of Ralstonia eutropha for the presence of phosphorylated residues during the PHB accumulation and PHB degradation phases. Thr373 of PHB synthase PhaC1 was phosphorylated during the stationary growth phase but was not modified during the exponential and PHB accumulation phases. Ser35 of PHB depolymerase PhaZa1 was identified in the phosphorylated form during both the exponential and stationary growth phases. Additional phosphosites were identified for both proteins in sample-dependent forms. Site-directed mutagenesis of the codon for Thr373 and other phosphosites of PhaC1 revealed a strong negative impact on PHB synthase activity. Modifications of Thr26 and Ser35 of PhaZa1 reduced the ability of R. eutropha to mobilize PHB in the stationary growth phase. Our results show that phosphorylation of PhaC1 and PhaZa1 can be important for the modulation of the activities of PHB synthase and PHB depolymerase.IMPORTANCE Poly(3-hydroxybutyrate) (PHB) and related polyhydroxyalkanoates (PHAs) are important intracellular carbon and energy storage compounds in many prokaryotes. The accumulation of PHB or PHAs increases the fitness of cells during periods of starvation and under other stress conditions. The simultaneous presence of PHB synthase (PhaC1) and PHB depolymerase (PhaZa1) on synthesized PHB granules in Ralstonia eutropha (alternative designation, Cupriavidus necator) was previously shown in several laboratories. These findings imply that the activities of PHB synthase and PHB depolymerase should be regulated to avoid a futile cycle of simultaneous synthesis and degradation of PHB. Here, we addressed this question by identifying the phosphorylation sites on PhaC1 and PhaZa1 and by site-directed mutagenesis of the identified residues. Furthermore, we conducted in vitro and in vivo analyses of PHB synthase activity and PHB contents.

From obtaining to degradation of PHB: A literature review. Part II

In our previous review (Part I - Material Properties), the chemical, physi-cal and morphological properties, applications and mechanisms of obtain-ing poly (3-hydroxybutyrate) were discussed. PHB is a biologically basedpolymer that presents itself as an ecological option due to its characteristicdegradation before environmental factors and due to the action of microor-ganisms such as algae, fungi and bacteria. In this second review, aspectsrelated to the types of degradation to which this polymer is subjectedand the necessary conditions for its degradation are addressed. Amongthese mechanisms are thermo-degradation, thermo-mechanical degrada-tion, abiotic degradation, photo degradation and biodegradation. It isunderstood that the degradation of PHB can be seen as an advantageousfeature of this material. This review complements the previous one anduses the same terminology.

Structure-property relationships in peroxide-assisted blends of poly(ε-caprolactone) and poly(3-hydroxybutyrate)

Poly(ε-caprolactone) and poly(3-hydroxybutyrate) (PCL/PHB) blends in two weight ratios (75/25 and 50/50) were reactively compatibilized in the presence of di-(2-tert-butyl-peroxyisopropyl)-benzene and dicumyl peroxide as free radical initiators. Rheological, mechanical, thermal properties and morphological features, as well as the chemical structure of PCL/PHB blends were investigated. It was found that regardless of PCL/PHB blend ratio, the viscosity of reactively compatibilized blends increased, approximately 4-fold for dicumyl peroxide and 7-fold for di-(2-tert-butyl-peroxyisopropyl)-benzene, which confirmed their partial branching/cross-linking. The results showed that studied peroxides are effective compatibilizers for PCL/PHB (75/25) blends. However, di-(tert-butylperoxyisopropyl)benzene peroxide was more efficient and its application in PCL/PHB (75/25) blends resulted in significant increase of the tensile yield strength and elongation at break of 38% and 144%, respectively. On the other hand, the compatibilization effects of peroxides on tensile properties of PCL/PHB (50/50) blends were negligible. This is due to free-radical induced degradation of PHB and formulation of low molecular weight compounds. These degradation products can act as plasticizers and reduce the interfacial tension in the phase boundary, which corresponded to changes in morphology and chemical structure of studied PCL/PHB blends.

Co-Expression of ORFCma with PHB Depolymerase (PhaZCma ) in Escherichia coli Induces Efficient Whole-Cell Biodegradation of Polyesters.

Whole-cell degradation of polyesters not only avoids the tedious process of enzyme separation, but also allows the degraded product to be reused as a carbon source. In this study, Escherichia coli BL21(DE3) harboring phaZCma , a gene encoding poly(3-hydroxybutyrate) (PHB) depolymerase from Caldimonas manganoxidans, is constructed. The extra-cellular fraction of E. coli/pPHAZ exhibits a fast PHB degradation rate where it only took 35 h to completely degrade PHB films, while C. manganoxidans takes 81 h to do the same. The co-expression of ORFCma (a putative periplasmic substrate binding protein that is within the same operon of phaZCma ) further improves the PHB degradation. While 28 h is needed for E. coli/pPHAZ to cause an 80% weight loss in PHB films, E. coli/pORFPHAZ needs only 21 h. Furthermore, it is able to degrade at-least four different polyesters, PHB, poly(lactic acid) (PLA), polycaprolactone (PCL), and poly(butylene succinate-co-adipate) (PBSA). Testing of the time course of 3-hydroxybutyrate concentration and the turbidity of the degradation solutions over time shows that PhaZCma has both exo- and endo-enzymatic activity. The whole-cell E. coli/pORFPHAZ can be used for recycling various polyesters while ORFCma can potentially be a universal element for enhancing the secretion of recombinant protein.

Thermal degradation of poly(caprolactone), poly(lactic acid), and poly(hydroxybutyrate) studied by TGA/FTIR and other analytical techniques

Thermal degradation of three biodegradable polyesters: poly(caprolactone), poly(lactic acid), and poly(hydroxybutyrate), was studied by thermogravimetry coupled to Fourier Transform Infrared Spectroscopy TGA/FTIR before and after they were partially degraded. TGA curves and Gram–Schmidt plots showed only one decomposition stage for both poly(caprolactone), PCL, and poly(lactic acid), PLA. In contrast, poly(hydroxybutyrate), PHB, exhibited two degradation stages by TGA, but only one region of evolved gases was appreciated in the Gram–Schmidt plot. It was established that hexenoic acid, e-caprolactone, and small fragments of polymeric chains are the main degradation products of PCL, which were simultaneously released during thermal decomposition of this polymer. Meanwhile, carboxylic acid, aldehydes, and lactide monomer and/or oligomers were evolved from degradation of PLA. Finally, carboxylic acids and ester moiety were detected in the course of degradation of PHB; thus, random chain scission reaction took place during thermal decomposition of this polymer. Results from the spectroscopic characterization (FTIR and 1H NMR) of partially degraded samples supported the degradation mechanisms suggested by TGA/FTIR studies.

From Obtaining to Degradation of PHB:Material Properties. Part I

This paper presents a review of the chemical, physical and morphological characteristics, as well as the existing applications and mechanisms forthe production of poly (3-hydroxybutyrate). This biopolymer, which isobtained from renewable sources, degrades when exposed in biologicallyactive environments and is biocompatible, that is, it is not rejected by thehuman body in health applications. However, in spite of presenting simi-lar properties with some conventional plastics, the PHB exhibits fragilebehavior and thermal instability when processed. The literature proposesthe use of blends, the development of copolymers or the insertion of addi-tives in an attempt to improve the mechanical and thermal properties of PHB.

The bifunctionality of poly[(R)-3-hydroxybutyrate] in self-reinforced composite materials

The poly[(R)-3-hydroxybutyrate] (PHB) is a highly crystalline, biosourced polymer. The advantages of the PHB are its biodegradability and biocompatibility; however, the brittleness caused by its high crystallinity decreases the application ability of the PHB in comparison with the polyolefins. Excellent results were observed for the reactive extrusion of PHB in the presence of peroxides in many investigations of the modifications of PHB mechanical properties. The disadvantage must be considered to be the thermal degradation of PHB during extended extrusion and its limitation in natural composite preparation. The peroxides are highly reactive with natural fillers, and this causes a decrease of the filler's mechanical properties. Consequently, the reactive extrusion may be a useful tool for the production of additives only. The results we present of this investigation is based on a different material preparation strategy. The two-stage method incorporated additives preparation via reactive extrusion of PHB and the blending of the obtained product with neat PHB. Theself-reinforced composite material obtained in this way revealed significantly higher values of stress and strain compared to neat PHB. The thermal degradation of the PHB matrix was retarded and total crystallinity of the composite was decreased. The materials were characterized using DSC, SEM and SEC techniques. The samples were also investigated employing tensile and impact strength tests.

The novel technique of vapor pressure analysis to monitor the enzymatic degradation of PHB by HPLC chromatography

A novel method was introduced for the quantitative determination of substances in aqueous solutions by using the evaporative light scattering (ELS) detector of a high performance liquid chromatograph (HPLC). The principle of the measurement is the different equilibrium vapor pressure of the solvent and the analyte resulting in decreasing evaporation rate, larger droplets and stronger signal with increasing concentration. The new technique based on vapor pressure analysis was validated with traditional UV-Vis detection carried out with a diode array detector (DAD). The new technique was used for monitoring the concentration of solutions obtained during the enzymatic degradation of poly(3-hydroxybutyrate) yielding the 3-hydroxybutyrate monomer as the product. The accuracy of the measurement allowed the determination of degradation kinetics as well. The results obtained with the two techniques showed excellent agreement at small concentrations. Deviations at larger concentrations were explained with the non-linear correlation between analyte concentration and detector signal and the linear regression used for calibration. Mathematical analysis of the method made possible the determination of the evaporation enthalpy of the analyte as well. The new approach is especially suitable for the quantitative analysis of compounds, which do not absorb in the detection range of the DAD detector or if their characteristic absorbance is close to the lower end of its wavelength range.