Branched-chain Alcohols Research Articles

Optimized synthesis of (Z)-3-hexen-1-yl caproate using germinated rapeseed lipase in organic solvent

Abstract (Z)-3-hexen-1-yl esters are important green top-note components of food flavors and fragrances. Effects of various process conditions on (Z)-3-hexen-1-yl caproate synthesis employing germinated rapeseed lipase acetone powder in organic solvent were investigated. Rapeseed lipase catalyzed ester formation more efficiently with non-polar compared to polar solvents despite high enzyme stability in both types of solvents. Maximum ester yield (90%) was obtained when 0.125 M (Z)-3-hexen-1-ol and caproic acid were reacted at 25 °C for 48 h in the presence of 50 g/L enzyme in heptane. Enzyme showed little sensitivity towards a w with optimum yield at 0.45, while added water did not affect ester yield. Esterification reduced by increasing molecular sieves (>0.0125%, w/v). The highest yields of caproic acid were obtained with isoamyl alcohol (93%) followed by butanol and (Z)-3-hexen-1-o1 (88%) respectively reflecting the enzyme specificity for straight and branched chain alcohols. Secondary alcohols showed low reactivity, while tertiary alcohol had either very low reactivity or not esterified at all. A good relationship has been found between ester synthesis and the solvent polarity (log P value); while no correlation for the effect of solvents on residual enzyme activity was observed. It may be concluded that germinated rapeseed lipase is a promising biocatalyst for the synthesis of valuable green flavor note compound. The enzyme also showed a wide range of temperature stability (5–50 °C).

Modelling of Processes of Decomposition of Organic Compounds of Municipal Solid Wastes in Cross Electromechanical Fields

At the present time, one of the most common methods of disposing the municipal solid wastes (MSW) is burial in sanitary landfills, where take place, long-term complex of physico-chemical and biochemical processes of decomposition. It was developed modeling of the natural process of decomposition of the solid Municipal wastes in cross electromechanical fields. The synthesized pellets was studied by IR spectroscopy. Residual hydrocarbons has a complex structure, represented by δ-lactones, cyclopentanone. The results of treatment of husk show there were the characteristic frequencies related to the OH group, indicating that the synthesis of dimers of branched chain alcohols (glycerides). Analysis of model shows that hydrocarbon product formed by the method of matching the external effects of the internal response of the reacting carbon-water composition.

Chemical and biochemical study of industrially produced San Simón da Costa smoked semi-hard cow's milk cheeses: Effects of storage under vacuum and different modified atmospheres

Two batches of smoked, semi-hard (ripened for 45 d) San Simón da Costa cow's milk cheeses with Protected Designation of Origin were used to investigate the chemical, biochemical, and sensorial parameters that may be affected by modified-atmosphere packaging. Cheeses were packaged for 45 d as follows: vacuum packaging, packaging in 100% N2, packaging in a gas mixture of 20% CO2/80% N2, and packaging in a gas mixture of 50% CO2/50% N2. The San Simón da Costa cheeses were characterized by high contents of lactic, oxalic, and citric organic acids. The main free amino acids found were isoleucine, phenylalanine, serine, valine, lysine, and glutamic acid, and the most abundant volatile compounds included ethanol, diacetyl, 2-butanol, isopropyl alcohol, furfural, acetaldehyde, 2-butanone, acetone, and 2-methylfuran. Modified atmospheres appeared to alter the ripening processes by affecting lipolysis, as indicated by the lower concentrations of butyric and propionic acids compared with control cheeses. In addition, modified-atmosphere packaging altered the proteolysis processes, yielding higher amounts of branched-chain alcohols. The results revealed that storage under modified atmosphere contributes to the accumulation of several compounds probably derived from smoke, including aldehydes such as 2-furancarboxaldehyde (furfural), alcohols such as 2-methoxyphenol (guaiacol), ketones such as 2-cyclopenten-1-one, and esters such as methyl furancarboxylate, which were negatively correlated with flavor. Vacuum packaging was the most useful technique in terms of preserving the sensory quality of San Simón da Costa Protected Designation of Origin cheeses. Considering the current demands for packaged portions of food at the distribution and retail levels and the potential health risks associated with some smoke-derived compounds usually present in some smoked foods, the results obtained in this study may be of special interest to the cheese industry.

Production of Dairy-Based, Natural Sulphur Flavor Concentrate by Yeast Fermentation

Production of sulphur flavor concentrate by yeast fermentation was investigated using dairy media (milk and cream) spiked with L-methionine. The yeasts studied included so-called dairy yeasts Candida kefyr, Kluyveromyces marxianus, Debaryomyces hansenii, Geotrichum candidum, and Yarrowia lipolytica as well as wine yeasts Saccharomyces cerevisiae and Saccharomyces bayanus. Methionol was produced as the predominant volatile sulphur flavor compound. Other volatile sulphur flavor compounds produced were dimethyl disulphide, S-methyl thioacetate, 3-methylthio-1-propanoic acid, methional, 3-methylthio-1-propene, and ethyl 3-methylthio-1-propanoate. In addition, numerous other volatile flavor compounds were produced, including ethanol, branched-chain alcohols, aromatic alcohol, ethyl esters, branched-chain esters, branched-chain fatty acids, and branched-chain aldehydes. The sensory attribute of a methionol sulphur flavor concentrate in cream is concentration-dependent, ranging from a savory-slight potato note to a strong potato-savory note to a cooked cheese-potato note.

Growth and aroma contribution of Microbacterium foliorum, Proteus vulgaris and Psychrobacter sp. during ripening in a cheese model medium

The growth and aroma contribution of Microbacterium foliorum, Proteus vulgaris and Psychrobacter sp., some common but rarely mentioned cheese bacteria, were investigated in a cheese model deacidified by Debaryomyces hansenii during the ripening process. Our results show that these bacteria had distinct growth and cheese flavour production patterns during the ripening process. P. vulgaris had the greatest capacity to produce not only the widest variety but also the highest quantities of volatile compounds with low olfactive thresholds, e.g. volatile sulphur compounds and branched-chain alcohols. Such compounds produced by P. vulgaris increased after 21 days of ripening and reached a maximum at 41 days. The three bacteria studied exhibited various degrees of caseinolytic, aminopeptidase and deaminase activities. Moreover, P. vulgaris had a greater capacity for hydrolysing casein and higher deaminase activity. Our results show that P. vulgaris, a Gram-negative bacterium naturally present on the surface of ripened cheeses, could produce high concentrations of flavour compounds from amino acid degradation during the ripening process. Its flavouring role in cheese cannot be neglected. Moreover, it could be a useful organism for producing natural flavours as dairy ingredients.

The addition of a cocktail of yeast species to Cantalet cheese changes bacterial survival and enhances aroma compound formation

Indigenous yeasts can be detected at high populations in raw milk Cantal cheese, a French Protected Denomination of Origin (PDO) hard cheese. To investigate their use as adjunct cultures to promote flavour development in Cantalet (small Cantal) cheese, three strains isolated from raw milk Cantal cheese, Kluyveromyces lactis, Yarrowia lipolytica, and Pichia fermentans were added at 3 (E3) and 5 (E5) log 10 colony-forming units (cfu)/mL to microfiltered milk at a ratio of 80/10/10 viable cells, respectively. The global microbial, compositional and biochemical changes induced by the presence of yeasts in cheese were determined. Adjunct yeasts did not grow but stayed at viable populations of approximately 4 and 6 log 10 cfu/g in E3 and E5 cheeses, respectively, throughout the ripening period. They were mainly constituted of K. lactis, while P. fermentans and Y. lipolytica were not detectable after 3 and 45 days of ripening, respectively. Several species of indigenous yeasts were also detected in E3 cheeses at the beginning of ripening only, and in the control cheeses without yeasts added. Lactoccoci survived for longer periods in the presence of yeast adjuncts, while, conversely, the viability of Streptococcus thermophilus decreased more rapidly. The addition of yeasts did not influence cheese composition and total free amino acid content. In contrast, it slightly increased lipolysis in both E3 and E5 cheeses and markedly enhanced the formation of some volatile aroma compounds. The concentrations of ethanol, ethyl esters and some branched-chain alcohols were 6 to 10 fold higher in E5 cheeses than in the control cheeses, and only slightly higher in E3 cheeses. This study shows that K. lactis has a potential as cheese adjunct culture in Cantalet cheese and that, added at populations of 4–5 log 10 cfu/g cheese, it enhances the formation of flavour compounds.

Flavouring Group Evaluation 62 (FGE.62) Consideration of linear and branched-chain aliphatic unsaturated, unconjugated alcohols, aldehydes, acids, and related esters evaluated by JECFA (61st meeting) structurally related to esters of branched- and straigh

Flavouring Group Evaluation 62 (FGE.62) Consideration of linear and branched-chain aliphatic unsaturated, unconjugated alcohols, aldehydes, acids, and related esters evaluated by JECFA (61st meeting) structurally related to esters of branched- and straigh

Non-fermentative pathways for synthesis of branched-chain higher alcohols as biofuels

Global energy and environmental problems have stimulated increased efforts towards synthesizing biofuels from renewable resources. Compared to the traditional biofuel, ethanol, higher alcohols offer advantages as gasoline substitutes because of their higher energy density and lower hygroscopicity. In addition, branched-chain alcohols have higher octane numbers compared with their straight-chain counterparts. However, these alcohols cannot be synthesized economically using native organisms. Here we present a metabolic engineering approach using Escherichia coli to produce higher alcohols including isobutanol, 1-butanol, 2-methyl-1-butanol, 3-methyl-1-butanol and 2-phenylethanol from glucose, a renewable carbon source. This strategy uses the host's highly active amino acid biosynthetic pathway and diverts its 2-keto acid intermediates for alcohol synthesis. In particular, we have achieved high-yield, high-specificity production of isobutanol from glucose. The strategy enables the exploration of biofuels beyond those naturally accumulated to high quantities in microbial fermentation.

Cold Flow Properties of Fatty Esters

This article is devoted to the study of cold flow properties of neat esters of branched chain alcohols with fatty acids and blends of these esters with fossil diesel fuel. According to determined CFPP values the influence of alcohol branching on the fuel filterability is negligible. Fossil fuel blending with fatty esters of branched alcohols up to 10vol % does not substantially change the cold flow properties of fossil fuel. The low-temperature properties of fossil diesel and fatty acid methyl/ethyl ester (FAME/FAEE) blends with low ester content (up to 10vol %), prepared mainly from oils/fats with higher share of saturated fatty acids namely palm oil, tallow and lard, were also measured. The obtained results show that studied esters do not change low-temperature properties of fossil fuel in the blends with the low esters content of 3–5vol %, although their own CFPP is over the value permissible by the standard EN 14 214. However, current standard EN 590 specifies to meet all parameters of the standard EN 14 214 also for FAME assigned for blended fuels entirely. The proposal to repeal the CFPP limit for esters assigned for blending is fully justified, without any negative effects on fuel quality.

GC–MS SPME profiling of rhizobacterial volatiles reveals prospective inducers of growth promotion and induced systemic resistance in plants

Chemical and plant growth studies of Bacilli strains GB03 and IN937a revealed that the volatile components 2,3-butanediol and acetoin trigger plant growth promotion in Arabidopsis. Differences in growth promotion when cytokinin-signaling mutants are exposed to GB03 versus IN937a volatiles suggest a divergence in chemical signaling for these two bacterial strains. To provide a comprehensive chemical profile of bacterial volatiles emitted from these biologically active strains, headspace solid phase microextraction (SPME) coupled with software extraction of overlapping GC-separated components was employed. Ten volatile metabolites already reported from GB03 and IN937a were identified as well as 28 compounds not previously characterized. Most of the newly identified compounds were branched-chain alcohols released from IN937a, at much higher levels than in GB03. Principal component analysis clearly separated GB03 from IN937a, with GB03 producing higher amounts of 3-methyl-1-butanol, 2-methyl-1-butanol and butane-1-methoxy-3-methyl. The branched-chain alcohols share a similar functional motif to that of 2,3-butanediol and may afford alternative structural patterns for elicitors from bacterial sources.

Assay for Sjögren–Larsson Syndrome Based on a Deficiency of Phytol Degradation

Sjogren–Larsson syndrome (SLS) is a metabolic disorder characterized by an accumulation of long-chain fatty alcohols in plasma (1)(2). Studies by Rizzo and coworkers (3)(4)(5) have identified the enzyme that is deficient in SLS as fatty aldehyde dehydrogenase (FALDH; EC 1.2.1.3), which is encoded by the ALDH10 gene. FALDH is part of the microsomal alcohol NAD+-oxidoreductase complex, which functions in the conversion of long-chain fatty alcohols into fatty acids (3). A deficiency of FALDH leads to the accumulation of fatty alcohols in plasma (4)(5). Interestingly, FALDH also plays a role in the degradation of leukotriene B4 (6). In our laboratory, we recently established that SLS patients are also deficient in the degradation of phytol (3,7,11,15-tetramethylhexadec-2-en-1-ol), a branched-chain fatty alcohol commonly found in nature as part of the chlorophyll molecule (7). Interest in phytol has focused mainly on its metabolism to phytanic acid, a fatty acid that plays an important role in Refsum disease and several other peroxisomal diseases (8)(9). However, very little was known about the precise mechanism of the conversion of phytol to phytanic acid until our discovery of the involvement of FALDH in this pathway. Indeed, when we incubated fibroblasts derived from SLS patients in the presence of phytol in the culture medium, we observed a marked deficiency in the conversion of phytol to phytenic acid (Fig. 1A⇓ ). In an effort to confirm this finding, we set up an assay in which fibroblast homogenates were incubated with phytol in the presence of NAD+ and assessed activity by quantifying the amount of phytenic acid formed in this reaction by gas chromatography–mass spectrometry (GC-MS) (7). This assay was both sensitive and reliable for the diagnosis of SLS patients, which is of interest because …

Synthesis of fatty acid esters from acid oils using lipase B from Candida antarctica

AbstractEsterification of corn and sunflower acid oils with straight‐ and branched‐chain alcohols were conducted using lipase B from Candida antarctica (Novozym 435) in n‐hexane. Sunflower acid oil consisted of 55.6% free fatty acids and 24.7% triacylglycerols, while the free fatty acids and triacylglycerols contents of corn acid oil were 75.3% and 8.6%, respectively. After 1.5 h of methanolysis of sunflower acid oil, the highest fatty acid methyl ester content (63.6%) was obtained at 40 °C and the total fatty acid/methanol molar ratio was 1/1, using 15% enzyme based on acid oil weight. The conversion of both acid oils with straight‐ and branched‐chain alcohols was not significantly affected by the chain length of the alcohols. However, the lowest fatty acid methyl ester content (50%) was obtained in the reaction of corn acid oil with methanol. Sunflower acid oil was converted to fatty acid esters using primer alcohols such as n‐propanol, i‐ and n‐butanol, n‐amylalcohols, n‐octanol, and a mixture of amylalcohol isomers, resulting in a fatty acid ester content of about 70% at 40 °C.

Evolution of the volatile components of ewes raw milk Zamorano cheese. Seasonal variation

The evolution during ripening and the seasonal variation of the volatile fraction of Zamorano cheeses were studied by using purge and trap extraction and GC-MS analysis. Concentrations of acetaldehyde, propanal and branched chain aldehydes, 2-methyl ketones, higher molecular weight alcohols, and most esters increased significantly with ripening time. Branched chain aldehydes were more abundant in spring and winter cheeses. 2-Heptanone showed lower concentrations in autumn cheeses, opposite to 2,3-butanedione and 2,3-pentanedione. Alcohols were in general lower in autumn cheeses, except ethanol and 2-propenol, which were lower in summer cheeses. Methyl-, propyl- and 2-butyl esters were in general less abundant in autumn cheeses. Discriminant analysis classified correctly 90% of the cheeses for the season of manufacture and 100% of the cheeses for the dairy of origin. Principal component analysis showed a positive correlation between odour intensity, aroma intensity, pungent taste and branched chain aldehydes, phenyl acetaldehyde and acetophenone. Odour and aroma qualities were negatively correlated with branched chain alcohols, butyl acetate and 2-butyl acetate.

Immobilized lipase-catalysed production of alkyl esters of restaurant grease as biodiesel.

Simple alkyl ester derivatives of restaurant grease were prepared using immobilized lipases as biocatalysts. The lipases studied included those of Thermomyces lanuginosa and Candida antarctica supported on granulated silica (gran- T.l. and gran- C.a., respectively), C. antarctica supported on a macroporous acrylic resin (SP435) and Pseudomonas cepacia immobilized within a phyllosilicate sol-gel matrix (IM PS-30). All alcoholysis reactions were carried out in solvent-free media employing a one-step addition of the alcohol to the reaction system. Of the lipases studied, IM PS-30 was found to be the most effective in catalysing the methanolysis and ethanolysis of grease. The processes catalysed by gran- T.l. and gran- C.a. lipases gave poor conversions to esters, and the SP435-catalysed reactions gave intermediate yields of ethyl and methyl esters. Water activity (a(w)) was an important factor in the methanolysis reactions; reaction media with a(w)<0.5 resulted in the highest conversions to methyl esters. Molecular sieves also improved methyl ester yields by as much as 20% in transesterification reactions catalysed by IM PS-30. The immobilized lipases also were evaluated for their ability to produce alkyl esters of grease with several additional normal and branched-chain alcohols.

Seasonal variation of volatile compounds in ewe raw milk La Serena cheese

La Serena cheese is a soft to semi-soft variety made in Extremadura (western Spain) from Merino ewe raw milk using an extract of Cynara cardunculus as milk coagulant. A purge and trap ap- paratus coupled to a GC-MS was used to study the seasonal variation of the volatile fraction of La Serena cheeses made at four artisan dairies. Alcohols and esters were the main chemical families found in La Serena cheese. Most volatile compounds were detected in cheeses made during the whole year, although at different concentrations. Abundances of hydrocarbons, primary alcohols, second- ary alcohols of an uneven number of carbon atoms, terpenes, 2,3-butanedione, free fatty acids, nitro- gen compounds and aromatic compounds were generally higher in cheeses made during spring. Aldehydes reached the highest levels in spring and summer cheeses. Ethanol and ethyl esters were more abundant in spring and autumn cheeses, while branched chain alcohols and branched chain alkyl esters were more abundant in summer cheeses. Cheeses made in winter showed a lower abun- dance of most volatile compounds, except dimethyl sulphide and dimethyl disulphide. Cheeses made in spring received the highest scores for odour and aroma quality, and cheeses made in winter re- ceived the lowest scores. High amounts of 2,3-butanedione, medium concentrations of most terpenes and alcohols, low levels of methyl ketones, especially of 2-butanone, and very low concentrations of 2-butanol were generally recorded for cheeses with the highest quality scores in sensory evaluation. Concentrations of most ethyl esters and branched chain alkyl esters in the highest scored cheeses were above the mean values in cheeses made during the whole year. However, levels of propyl esters and ethyl esters of branched chain acids were lower in the highest scored cheeses. Volatile compound / ewe raw milk / La Serena cheese / seasonal variation

Evolution of the volatile components of ewe raw milk La Serena cheese during ripening. Correlation with flavour characteristics

La Serena cheese is a soft to semi-soft Spanish variety manufactured from Merino ewe raw milk coagulated with an extract of Cynara cardunculus. In the present work a purge and trap ex- traction procedure coupled to gas chromatography - mass spectrometry was used to study the volatile fraction evolution during ripening of La Serena cheese. A total of 112 compounds were identified. Al- cohols were quantitatively the main chemical family in the volatile fraction, increasing significantly during ripening. Ethanol, 1-propanol, 2-propenol, 2-propanol, 2-butanol, 2-pentanol and the branched chain 2-methyl-1-propanol and 3-methyl-1-butanol, were abundant at the end of ripening. Esters, especially ethyl esters of acetic, butanoic, hexanoic and octanoic acids and 3-methyl-1- butanol acetate, were found at high levels in the headspace of La Serena cheese. Concentrations of most esters increased dramatically during ripening, and, given their low perception thresholds, may be considered as key constituents of the aroma of this cheese variety. The principal component analy- sis of sensory attributes and volatile compounds extracted 3 functions. Linear aldehydes and alco- hols, 2-alkanols, branched-chain alcohols, 2-methyl-ketones, alkanes, terpenes and esters other than ethyl esters correlated positively with function 1 (25.8% of the variance explained). Odour and aroma quality, along with the sensory attributes lactic, fruits-flowers, and clean cheese flavour, and the volatiles ethanol and ethyl esters, had a high correlation coefficient with function 2 (14.8% of the vari- ance explained). 2-Butanone and the sensory attributes fishy and putrid had a negative correlation co- efficient with this function. Odour and aroma intensity, along with the sensory attributes animal, pungent and fishy, and the volatile compounds branched chain aldehydes, n-alkanols, 2-butanol and esters, were positively correlated with function 3 (11.0% of the variance explained). Terpenes had a negative correlation coefficient with this function. Volatile compound / ewe raw milk / La Serena cheese / ripening / flavour

Extremely stable and versatile carboxylesterase from a hyperthermophilic archaeon.

We have found that the hyperthermophilic archaeon Pyrobaculum calidifontis VA1 produced a thermostable esterase. We isolated and sequenced the esterase gene (est(Pc)) from strain VA1. est(Pc) consisted of 939 bp, corresponding to 313 amino acid residues with a molecular mass of 34,354 Da. As est(Pc) showed significant identity (30%) to mammalian hormone-sensitive lipases (HSLs), esterase of P. calidifontis (Est) could be regarded as a new member of the HSL family. Activity levels of the enzyme were comparable or higher than those of previously reported enzymes not only at high temperature (6,410 U/mg at 90 degrees C), but also at ambient temperature (1,050 U/mg at 30 degrees C). The enzyme displayed extremely high thermostability and was also stable after incubation with various water-miscible organic solvents at a concentration of 80%. The enzyme also exhibited activity in the presence of organic solvents. Est of P. calidifontis showed higher hydrolytic activity towards esters with short to medium chains, with p-nitrophenyl caproate (C(6)) the best substrate among the p-nitrophenyl esters examined. As for the alcoholic moiety, the enzyme displayed esterase activity towards esters with both straight- and branched-chain alcohols. Most surprisingly, we found that this Est enzyme hydrolyzed the tertiary alcohol ester tert-butyl acetate, a feature very rare among previously reported lipolytic enzymes. The extreme stability against heat and organic solvents, along with its activity towards a tertiary alcohol ester, indicates a high potential for the Est of P. calidifontis in future applications.

Phase Transition in Monolayers of Straight Chain and 2-Methyl Branched Alcohols at the Air−Water Interface

Monolayers of straight chain and 2-methyl branched chain alcohols with alkyl chain lengths of C10−C18 are experimentally studied by a conventional film balance technique combined with a Brewster angle microscope (BAM). The comparison of the surface pressure (π)−area (A) isotherms with the corresponding BAM images provides information on the phase behavior and the first-order main phase transition of the monolayers. Striking differences in the dependence of the phase transition pressure on temperature of the straight chain and 2-methyl-substituted alcohols are correlated with differences in the molecular ordering. The general conditions for the main phase transition in the corresponding homologous alcohols can be derived. The effect of alkyl chain length and 2-methyl substitution on the general textural features of the condensed phase domains is determined under equilibrium and nonequilibrium conditions. n-Alcohol monolayers form defined and well-shaped condensed phase domains, often with inner texture in ...

Genetic and physiological analysis of branched-chain alcohols and isoamyl acetate production in Saccharomyces cerevisiae

Branched-chain alcohols, such as isoamyl alcohol and isobutanol, and isoamyl acetate are important flavor components of yeast-fermented alcoholic beverages. Analysis of a null mutant of the BAT2 gene encoding cytosolic branched-chain amino acid aminotransferase, and a transformant with multi-copy plasmids containing the BAT2 gene showed that the BAT2 gene product plays an important role in the production of branched-chain alcohols and isoamyl acetate. Fermentation tests using the bat2 null mutant transformed with multi-copy plasmids carrying the ATF1 gene, which encodes alcohol acetyltransferase, indicated that modified expression of BAT2 and ATF1 genes could significantly alter the proportion of branched-chain alcohols and isoamyl acetate synthesized. Furthermore, fermentation tests using different ratios of nitrogen source and RNA blot analyses demonstrated that transcription of L-leucine biosynthetic ( LEU) and BAT genes is co-regulated by nitrogen source, that production of isoamyl alcohol depends on this transcription, and that ATF transcription increased with increased concentrations of nitrogen source. Our data suggest that changes in isoamyl alcohol production by nitrogen source are due to transcriptional co-regulation of LEU and BAT genes, and that production of isoamyl acetate is dependent on isoamyl alcohol production and ATF transcription.

Effects on male rats of di-(2-ethylhexyl) phthalate and di- n-hexylphthalate administered alone or in combination

The effects of phthalate esters of branched chain alcohols, typified by di-(2-ethylhexyl)phthalate (DEHP) differ from those of esters of straight chain alcohols typified by di -n-hexyl phthalate (DnHP). The former induce liver enlargement and proliferation of hepatic peroxisomes, while the latter cause no peroxisome proliferation but cause fat accumulation in the liver. Both classes of phthalate esters are hypolipidaemic and cause thyroid changes associated with an increased rate of thyroglobulin turnover. As phthalate esters are used as mixtures, we have examined the effect of mixtures of the compounds. Groups of five male Wistar albino rats were administered either control diet or diets containing either 10 000 ppm of DEHP, 10 000 ppm of DnHP or 10 000 ppm DEHP plus 10 000 ppm DnHP for 14 days. Rats receiving diets containing DEHP showed the expected increase in relative liver weight, in “peroxisomal” fatty acid oxidation and in CYP4A1. Serum triglyceride and serum cholesterol were also reduced, and the thyroid showed the histological changes mentioned above. Rats consuming diets containing DnHP showed no increase in relative liver weight and no induction of peroxisomal fatty acid oxidation or CYP4A1. However, there was a marked accumulation of fat in the liver. The fall in serum cholesterol was similar to that in rats treated with DEHP, but the fall of serum triglyceride was more pronounced. Thyroidal changes were again observed. In general, changes in rats treated with a mixture of DEHP and DnHP were very similar to those found with rats treated with DEHP alone. The liver was enlarged, and peroxisomal fatty acid oxidation and CYP4A1 were both induced. The amount of fat in the liver was much less than in rats receiving DnHP alone. Thyroid changes were similar to those in rats receiving the individual compounds. The effect on serum cholesterol seemed additive, but the levels of serum triglyceride were intermediate between the groups receiving the single compounds.