Heritability of polyphenols, anthocyanins and antioxidant capacity of Cameroonian cocoa (Theobroma cacao L.) beans

This study investigates the heritability of polyphenolic, anthocyanin and antioxidant capacity of beans derived from four cocoa clones and their offsprings. These compounds were analyzed at 280 nm (polyphenols) and 520 nm (anthocyanins) by reversed-phase high-performance liquid chromatography (RP-HPLC) method using a photodiode array detector (PDA). The antioxidant capacity of methanolic extracts prepared from cocoa beans was measured by the DPPH and ABTS assays. Samples displayed catechin and epicatechin as the two main polyphenols. Epicatechin, which represents 1 to 5% of defatted cocoa seed powder, was hundred-fold higher than catechin while cyanidin-3-arabinoside was three-fold higher than cyanidin-3-galactoside. The two main anthocyanins found in our samples represent about 0.05% of defatted cocoa seed powder. All these compounds were genotype-dependent. Unidentified substances called A, B and C were also found in cocoa seeds. Substance A is discussed as a derivative of caffeic acid and an ester-bound compound. Substances B and C are oligomers of proanthocyanidins. Antioxidant capacity of cocoa beans obtained by 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid (ABTS) assay was higher than those obtained by DPPH assay. According to correlation tests, flavanols contributed better in the antioxidant capacity than anthocyanin. No maternal effect was detected in the transmission of polyphenol compounds suggesting a nuclear heritability.Key words: Theobroma cacao, cocoa beans, polyphenolic compounds, anthocyanins, heritability

Change in amino acids content during germination and seedling growth of Cola sp.

Accessions of Cola acuminata and Cola anomala were analysed based on the variation of free amino acids during germination and seedling growth. These compounds were determined by high performance liquid chromatography (HPLC) methods. In the non-germinated seeds of C. acuminata, arginine (Arg) was the major amino acid found in all six accessions and valine (Val) was the least present, while in C. anomala, gamma ()-aminobutyric acid (GABA) was predominant and isoleucine (Ile) was less abundant. During germination, the seeds of the two species accumulated large amount of GABA and very little tryptophan. Principal component and cluster analyses of Cola sp. categorized the accessions into two groups distinguishing the two Cola species. The differences between the groups were related not only to the absence/presence of histidine (His) and phenylalanine (Phe), but also to amide amino acids asparagine (Asn), glutamine (Gln) and Arg contents. In Cola sp., free amino acids varied significantly during these two processes indicating their high utilization. These metabolites can be considered as crucial in the control of germination.Keywords: Cola sp., amino acids, germination, seedling growthAfrican Journal of Biotechnology Vol. 9(35), pp. 5632-5642, 30 August, 201

Sulphur depletion altered somatic embryogenesis in Theobroma cacao L. Biochemical difference related to sulphur metabolism between embryogenic and non embryogenic calli

Somatic embryogenesis is a useful tool for Theobroma cacao improvement and propagation. Depending on culture medium composition, different morphogenetic structures (including somatic embryo) occur in response to alteration of genes expression patterns and biochemical changes. The effect of SO42- ion deficiency in culture media on somatic embryogenesis was studied through sequential replacement of MgSO4 and K2SO4 by MgCl2 and KCl, respectively, at different steps of somatic embryogenesis. It appears that explants gradually lost their embryogenic competence as the period of exposition to sulphur free medium increases. These results suggest that, sulphur availability and the duration to sulphur exposition might modulate the expression of genes involved in somatic embryo differentiation in T. cacao. Cysteine, glutathione, reducing sugars, cysteine synthase and cysteine desulfurase activities were analysed in different morphogenetic structures obtained in vitro. Cysteine and reducing sugars contents appeared to be higher in embryogenic calli than their nonembryogenic homologues, whereas glutathione content appears to be lower in embryogenic calli. Cysteine synthase activities also discriminate the embryogenic calli from non embryogenic calli. In the embryogenic calli, the ratio cysteine synthase/cysteine desulfurase activities were above unit. The assimilation of exogenous sulphur (sulphate) for the synthesis of cysteine might hence be crucial for somatic embryogenesis in T. cacao. This explains the reduction and the absence of somatic embryo response observed during sulphur depletion in culture media. Sulphur nutrition is therefore critical in cacao somatic embryogenesis.Keyswords: Cacao, embryo, sulphate, cysteine synthase, glutathione, deficiencyAfrican Journal of Biotechnology Vol. 9(35), pp. 5665-5675, 30 August, 201

The Influence of fermentation-like incubation on cacao seed testa and composition of testa associated mucilage

Because the testa of Theobroma cacao L. is a physical barrier for mass transfer of water and nutrients during germination, as well as in the fermentation process, analysis of the testa is necessary to understand these processes. This research deals with the morphological structure of the testa and its influence during fermentation of cacao seeds on the production of the aroma precursors of chocolate. Investigations have been carried out using fermentation-like incubations. For the first 48 hours of fermentation-like incubations, the seed represents a closed system with no transport of matter between the incubation-medium outside and the cotyledons inside. At this stage, the testa influences the course of the incubations by its barrier function. Much of this activity involves testa-associated mucilage. The composition of testa-associated mucilage is described

German cacao of Cameroon - new facts on a traditional variety fallen into oblivion

“German” cacao cultivated in Cameroon has emerged from a mixture of different gene pools with a large proportion of Trinitario and with a pronounced content of polyphenols. In order to characterize this old genotype, polyphenols and polyphenol oxidase were compared with hybrid selected genotypes. Epicatechin (25 mg/g – 52 mg/g ffdm) and catechin (0.5 – 1.9 mg/g ffdm) content of German cacao seeds were of similar range with hybrid investigated samples. German cacao is characterized by its high content of anthocyanins especially cyadinine-3-arabinosid with a range from 8.84 mg/g to 17.51 mg/g ffdm. Hybrid genotypes displayed 1 mg/g to 6.4 mg/g ffdm of cyanidine-3-arabinosid. PPO activity was 10 to 20-fold higher in German cacao seeds compared to hybrid. Anthocyanin and PPO through the oxidation of phenols to quinone are involved in colour development and pests and diseases resistance. Pigment is one of the most important factors for the colour of cocoa powder. We discussed the high content of anthocyanin and PPO activity in German cacao in relation with the reddish colour of cocoa powder derived from Cameroonian cacao

HETEROSIS AND HERITABILITY ESTIMATES OF PURINE ALKALOIDS AND POLYPHENOLS IN COCOA

Cocoa ( Theobrama cacao L. ) is an important allogamous tropical tree crop, whose centre of diversity is considered to be in Central America. Dry cocoa beans from five cocoa clones, and their intercrossed hybrids were analysed based on the variation of alkaloids and polyphenolic compounds contents, in order to gain insights on the heterosis and broad-sense heritability. Polyphenols and alkaloids were analysed at 280 nm by HPLC, using a Photodiode Array Detector (PDA); while anthocyanins were separated with the SEP-PAK Vac 6cc 1000 mg (waters) column and measured at 520 nm with a PDA. Dry cocoa beans displayed high content of purine alkaloids (2.1 and 8.8 mg g-1 for caffein and theobromine, respectively), and polyphenols (25 and 2978 \ub5g g-1 for catechin and epicatechin, respectively). Among the five cocoa clones, SNK16 was the highest in purine alkaloid (caffein and theobromin) and flavanol (catechin and epicatechin); while T79/467 possessed the greatest quantity of cyanidin-3-galactoside and cyanidin-3-arabinoside. From all the parameters studied, anthocyanins (Cyanidin-3-galactoside and cyanidin-3-arabinoside) exhibited the highest level of heterosis. Parental genotypes SNK16 and T79/467 showed good aptitudes for the combination of characters because their reciprocal hybrids F5 and F9, distinguished themselves by better levels of mid-parent heterosis values. Besides, the heritability value in strict sense of this Cyanidin-3-galactoside was very high. Absence of significant difference between genotypes, coming from reciprocal crossbreeding for Cyanidin-3-galactoside, suggests that this character in cocoa would be nuclear contrary to purine alkaloids and flavan-3-ols, where their transmission to offsprings can be stated as cytoplasmic.Le cacaoyer ( Theobrama cacao L. ) est une importante plante tropicale allogame originaire d\u2019Am\ue9rique Centrale. Les teneurs en alcalo\uefdes et en polyph\ue9nols ont \ue9t\ue9 analys\ue9es sur des f\ue8ves de cinq clones de cacao et de leurs descendants issus des croisements r\ue9ciproques afin de d\ue9terminer l\u2019h\ue9t\ue9rosis et l\u2019h\ue9ritabilit\ue9 de ces m\ue9tabolites. Les alcalo\uefdes et les polyph\ue9nols ont \ue9t\ue9 analys\ue9s par HPLC \ue0 280nm utilisant un d\ue9tecteur \ue0 barrettes de photodiode (PDA) alors que les anthocyanines l\u2019ont \ue9t\ue9 sur une colonne SEP-PAK Vac 6cc 1000 mg \ue0 520nm utilisant le PDA. Ces f\ue8ves ont des teneurs \ue9lev\ue9es en alcalo\uefdes puriques (2,1 et 8,8 mg.g-1 de caf\ue9ine et de th\ue9obromine respectivement) et en polyph\ue9nols (25 \ub5g.g-1 de cat\ue9chine et 2978 \ub5g.g-1 d\u2019epicat\ue9chine). Des cinq clones utilis\ue9s, SNK16 s\u2019est distingu\ue9 par des teneurs les plus \ue9lev\ue9es en caf\ue9ine, th\ue9obromine, cat\ue9chine et \ue9picat\ue9chine. De tous les param\ue8tres analys\ue9s, les anthocyanines ont montr\ue9 un niveau d\u2019h\ue9t\ue9rosis \ue9lev\ue9. Les g\ue9notypes SNK16 et T79/467 ont pr\ue9sent\ue9 une meilleure aptitude \ue0 la combinaison et les hybrides issus de leur croisement r\ue9ciproque (F5 et F9) ont pr\ue9sent\ue9 une meilleure h\ue9t\ue9rosis par rapport au meilleur parent. L\u2019utilisation de ces deux clones dans un champ semencier serait tr\ue8s importante pour des industries pharmaceutiques car leur croisement g\ue9n\ue8re des hybrides \ue0 haut potentiel en alcalo\uefdes et en polyph\ue9nols. De plus, l\u2019absence d\u2019une diff\ue9rence significative de cyanidine-3-galactoside entre hybrides r\ue9ciproques sugg\ue8re que l\u2019h\ue9ritabilit\ue9 de ce caract\ue8re serait nucl\ue9aire contrairement aux alcalo\uefdes puriques et aux flavan-3-ols dont l\u2019h\ue9ritabilit\ue9 serait de nature cytoplasmique

Biochemical profile of cuttings used for in vitro organogenesis of Prunus africana: An endangered species in Cameroon

Prunus africana (Kanda stick) is a valuable medicinal tree classified as priority species for domestication in Cameroon. To measure the ability for developing axillary buds and multiple shoots in relation with the origin and state of explants, some biochemical parameters were assessed in eight types of single-node cuttings. These cuttings were treated with different concentrations of kinetin (kin) indole butyric acid (IBA) and thidiazuron (TDZ) and cultured in Murashige and Skoog (MS) medium. The glutelin content (1.13 ± 0.201 µg/mg FW) was higher compared to those of total albumin, globulin and prolamin and most representative at mature vegetative apical stage (MVA). The highest (15.60 ± 1.966 µg/mg FW) and lower (2.82 ± 0.072 µg/mg FW) amount of phenol compound was observed in the mature dormant stages and immature vegetative distal stage (IVD), respectively. Regarding the peroxidases, highest activity (0.453 ± 0.055 OD/mg/ FW.5 min-1) was obtained in mature dormant apical stage. Apart from IVA state, there was no reactive explant cultured in vitro. The high percentage of budding and proliferation of budding (100%) and rooting (100%) were achieved with 12 μM Kin and 24 μM IBA, respectively. In the presence of 24 μM TDZ, multiple shoot was induced with a maximum of 5 shoots/explants. Micropropagation success of P. africana is influenced by cytokinin, physiological and biochemical status of single node cutting.Key words: Biochemical profile, Prunus africana, budding, multiple shoots, rooting

Dynamics of Auxin and Cytokinin Metabolism during Early Root and Hypocotyl Growth in Theobroma cacao

The spatial location and timing of plant developmental events are largely regulated by the well balanced effects of auxin and cytokinin phytohormone interplay. Together with transport, localized metabolism regulates the concentration gradients of their bioactive forms, ultimately eliciting growth responses. In order to explore the dynamics of auxin and cytokinin metabolism during early seedling growth in Theobroma cacao (cacao), we have performed auxin and cytokinin metabolite profiling in hypocotyls and root developmental sections at different times by using ultra-high-performance liquid chromatography-electrospray tandem mass spectrometry (UHPLC-MS/MS). Our work provides quantitative characterization of auxin and cytokinin metabolites throughout early root and hypocotyl development and identifies common and distinctive features of auxin and cytokinin metabolism during cacao seedling development

Assessment of the profile of free amino acids and reducing sugars of cacao beans from local Cameroonian Trinitario (SNK varieties) and Forastero (TIKO varieties) using fermentation-like incubation

This study investigated the profile of cacao beans from the local Cameroonian Trinitario (SNK) and Forastero (TIKO) in terms of aroma precursors (amino acids and reducing sugars) through fermentation-like incubations. Treatments consisted of incubating beans in acetic acid, 100 mmol/L for two days followed by 200 mmol/L of acetic acid for three days (Treatment T1) and in 100 mmol/L of lactic acid for two days followed by 200 mmol/L of acetic acid for three days (Treatment T2). Both treatments resulted in an increase of free amino acids content by 1.5 - 2.5 times in SNK and TIKO varieties. The ratio of the hydrophobic amino acids over the rest of amino acids showed the preponderance of T1 on the hydrophobic amino acids released in TIKO while in the SNK, some varieties displayed the highest ratio in T2. Glucose and fructose content in TIKO and SNK beans increased 2 to 3 times during incubation. Galactose and raf-finose were found in unfermented beans. After incubation, raffinose was missing while at the same time a raise of galactose content could be seen. These results highlighted that acidification remains the factor inducing the releasing of free hydrophobic amino acids, the genotype being less involved